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The Pacific white shrimp Penaeus vannamei is one of the most productive shrimp species in China and the world. Domestic demand for P. vannamei has increased annually because of its fast growth, easy breeding, strong resistance to stress and disease, and high meat yield. The primary challenge in the development of the shrimp industry is seed rearing. The demand for broodstocks of P. vannamei in China is as high as 1, 000, 000 pairs per year, and the total weight of shrimp seedlings exceeds 1.3 trillion. China needs to import 300, 000 pairs of P. vannamei every year. The cost of each pair of P. vannamei is approximately 200 US＄, thus equating to hundreds of millions of US＄ in total. During the production of P. vannamei, the spawning frequency varies greatly among female shrimp. Cultivating high-fecundity P. vannamei through genetic improvement is an effective way to reduce the cost of seedling farms and improve economic benefits.To further study the molecular mechanisms of fecundity differences in female P. vannamei, the ovarian histology and expression of four candidate genes in populations with high- and low-fecundity were analyzed in this study. Before the experiment, the 7-month-old parent shrimp of P. vannamei were fed a commercial parent shrimp fortified diet, a maturation promoting diet, and squid for nutritional enhancement. After one month of promoting maturity, the unilateral eyestalk of female shrimp was cut using forceps, and next-generation family construction began after 15–20 days of recovery. The spawning frequency of 439 female shrimp in the 30-day production cycle was calculated and used to indicate fecundity. Those with a spawning frequency of 0 were included in the low-fecundity group, while those with a spawning frequency of > 3 were included in the high-fecundity group. The ovarian development of P. vannamei was divided into four stages (proliferation, small growth, large growth, and maturation) according to the color, size, and shape of the ovaries. In the high-fecundity group, the ovaries exhibited development up to stageⅣ, indicating their attainment of maturity. In the low-fecundity group, ovarian development was limited to stageⅢ, with progression to stageⅣ proving difficult. Ovarian tissues from the same part of female shrimp of stageⅠ–Ⅳ in the high-fecundity group and stageⅠ–Ⅲ in the low-fecundity group were taken and immediately immersed in tissue fixative for paraffin sectioning. The research team previously identified candidate genes related to fecundity in genomic regions undergoing selective sweep in isolated populations. These included thioredoxin 2 (TRX2), partitioning-defective 3 (PARD3), phospholipase Cβ4 (PLCβ4), and arginine-glutamate acid dipeptide repeats (RERE), which play important roles in oocyte maturation, cell proliferation, and early development in other species. qRT-PCR was used to analyze and compare their expression in the ovary and eyestalk tissues of high- and low-fecundity females at different stages of ovarian development.The results of paraffin section showed that the ovarian development of the low-fecundity group was slower than that of the high-fecundity group. In the high-fecundity group, stageⅠ ovaries primarily consisted of oogonia and oocytes during the early phase of yolk formation, with the emergence of small spherical yolk granules. StageⅠ ovaries in the low-fecundity group primarily consisted of oogonia, with no observable yolk granule formation. StageⅡ ovaries in the high-fecundity group were mainly composed of oocytes with vitelline formation, accompanied by a notable augmentation in both the size and number of yolk granules. StageⅡ ovaries in the low-fecundity group primarily comprised oocytes in the early phase of vitelline formation with yolk granules beginning to form. StageⅢ ovaries in the high-fecundity group consisted mainly of oocytes during the late phase of yolk formation. Many spherical yolk granules were formed in the cytoplasm, and cortical rods began to appear, although the lengths of the cortical rods were short. StageⅢ ovaries in the low-fecundity group were composed of oocytes during yolk formation. The cytoplasm was full of spherical yolk granules in the absence of cortical rods. In the high-fecundity group, stage Ⅳ ovaries were filled with mature oocytes, large yolk granules filled the entire cytoplasm, and thick cortical rods were formed in the oocytes. qRT-PCR showed that the four genes maintained high expression levels in the mature ovaries of the high-fecundity group. Their expression levels in the ovaries of the high-fecundity group were higher than those in the ovaries of the low-fecundity group in the small and large growth stages. TRX2, PLCβ4, and RERE expression levels were significantly different between the groups (P < 0.05). In eyestalk tissues at different stages, TRX2, PARD3, PLCβ4, and RERE expression levels in the low-fecundity group were higher than those in the high-fecundity group. The results suggest that these four genes may play important roles in the ovarian development of P. vannamei. The expression levels of all four genes in the high-fecundity group were lower than those in the low-fecundity group in eyestalk tissues. However, in ovarian tissues, except for the PARD3 gene in stageⅠ, which was expressed at lower levels in the high-fecundity group than in the low-fecundity group, the expression levels of all four genes in the other stages were higher in the high-fecundity group than those in the low-fecundity group, which is opposite to the expression trend in eyestalk tissues. Therefore, we speculate that these genes also promote cell proliferation, development, and target gene expression in the eyestalks of the low-fecundity group. Their high expression in the eyestalks of the low-fecundity group may enhance the secretion of ovarian inhibitory factors, thereby inhibiting ovarian development. Further research is needed to reveal their functional mechanisms. The above results provide an important reference for the in-depth study of the molecular mechanisms of differences in the fecundity of female P. vannamei.

Penaeus vannamei, also known as white foot shrimp, is globally one of the three high-yield shrimp farming varieties. The wild species is found along the Pacific coast of South America. Since 1988, China has introduced P. vannamei. Due to its strong environmental adaptability, high feed conversion rate, fast growth rate, and high tolerance to ammonia nitrogen and nitrite, it has been widely promoted in aquaculture. By 2021, the aquaculture output of P. vannamei in China reached 1.977 million tons, accounting for approximately 37% of the world's total production, and has extremely high economic value.Growth traits are among the most important economic factors in the production of P. vannamei. With the intensive development of shrimp farming and degradation of germplasm, the white spot syndrome virus (WSSV) is a serious disease faced by the global shrimp industry. The infection can cause symptoms such as reduced food intake, enlarged liver and pancreas, pale red body color, and white spots on the head and chest armor in shrimp, resulting in widespread death.At present, China has cultivated 12 new varieties of P. vannamei, which to some extent alleviates its dependence on imports for high-quality P. vannamei. However, the excellent traits of domestic shrimp species are singular, and cultivating new varieties of P. vannamei with fast growth and strong disease resistance is an urgent demand in the market. Genetic parameter evaluation is the fundamental work of selection breeding. Heritability reflects the genetic variation in traits in the breeding population, which is of great significance in the development of a selection index, prediction of selection response, comparison of selection methods, selection breeding planning, and other breeding processes. There are different degrees of genetic correlation among various quantitative traits of shrimp, and genetic correlation coefficients are important for selecting target traits. The estimation of genetic correlation can be used to develop a comprehensive selection index and breeding program of multiple traits, which can improve the selection efficiency and breed better varieties with multiple traits. The higher the genetic correlation between traits, the better the effect of indirect selection. The estimation of genetic parameters is greatly affected by the test population, breeding management, analysis methods, and other factors. To ensure the accuracy of multi-trait composite breeding for growth and WSSV resistance, accurate evaluation of growth and WSSV resistance needs to be carried out for specific breeding populations. There are two commonly used target traits for measuring WSSV resistance in P. vannamei: individual survival time after infection and half-lethal survival rate (SS50) of families. In practice, measuring the half-lethal survival rate of families is more convenient. However, the correlation between these two traits has not yet been reported.To promote the growth and WSSV resistance of P. vannamei, 59 families (1, 770 individuals) were tested for WSSV infection. We recorded the survival time and individual body length of shrimp after infection and analyzed the mean, standard deviation, maximum and minimum values, and coefficient of variation of half-lethal survival rate and test traits for each line. The heritability and genetic correlation coefficients of growth, survival time, and half-lethal survival rate were calculated. Variance components and genetic parameters for growth and survival traits were estimated using a two-trait animal model and a sire-dam threshold model. The genetic parameters of body length were corrected by including age as a covariate. The genetic parameters of WSSV survival time were corrected by including body length as a covariate. The estimated heritability of body length was medium (0.17±0.05), and the estimated heritabilities of survival time and half-lethal survival rate were medium (0.18±0.05 and 0.14±0.05). Further, the estimated heritabilities of the three traits were significantly different from zero (P < 0.01). The genetic correlation between body length and survival time and that of body length and half-lethal survival rate were low (0.15±0.20 and 0.25±0.22). There were no significant differences between the genetic correlations and zero (P > 0.05). The genetic correlation between survival time and half-lethal survival rate was high (0.96±0.03). There was no significant difference between the genetic correlation and one (P > 0.05). The results showed that a comprehensive selection index of growth and WSSV resistance of P. vannamei can be established to carry out multi-trait composite breeding. In this breeding population, the growth and WSSV resistance of P. vannamei can be combined with multiple traits according to the breeding requirements, and the comprehensive selection index can be formulated by assigning values. In addition, to optimize the breeding of each generation, the half-lethal survival rate of WSSV can be used as an indicator of WSSV resistance. This study provides basic data and theoretical support for breeding varieties of P. vannamei with excellent growth and WSSV resistance.

Pacific white shrimp Penaeus vannamei, as one of the most productive and economically valuable shrimp species in the world, have many favorable characteristics, such as a short growth cycle, strong adaptability and disease resistance, and a low nutritional demand. They are omnivorous animals whose diet mainly consists of animal-derived food. Conventional feed used in their culture contains fishmeal as the main protein source. Feed costs account for 40%~60% of their production costs. Therefore, breeding P. vannamei strains without demand for fishmeal would be an effective way to address the issues of fishmeal shortage and high feed costs. Improving utilization efficiency for diets without fishmeal is key to breeding such P. vannamei strains. In this study, genetic parameters for feed efficiency traits (feed efficiency ratio, feed intake, and weight gain) and genotype by environment interaction were evaluated using two diets, one without fishmeal (38% protein) and one with 25% fishmeal (42% protein).Thirty families of P. vannamei were selected for a 40-day feeding experiment. A total of 900 shrimp (30 animals per family) were selected. Fifteen animals per family were fed a zero-fishmeal diet, and the other 15 animals were fed a high fishmeal diet. The feed intake and body weight of all animals were measured for calculating the feed efficiency ratio and weight gain. The variance components for these traits were estimated using a univariate model and an average information restricted maximum likelihood algorithm, and genetic (phenotypic) correlations between traits were estimated using bivariate analysis.The results showed that shrimp fed a high fishmeal diet exhibited a significantly higher mean weight gain (62.00%) and feed efficiency ratio (124.00%) than those fed a zero-fishmeal diet, which had a mean weight gain of 23.50% and feed efficiency ratio of 49.40%. There was a small difference in the mean feed intake between shrimp fed a high fishmeal diet (7.09 g) and those fed a zero-fishmeal diet (7.28 g).The heritability estimates for feed intake of shrimp fed zero- and high fishmeal diets were 0.699 ±0.155 and 0.556±0.139, respectively. For weight gain, the heritability estimates of shrimp fed zero- and high fishmeal diets were 0.458±0.140 and 0.667±0.146, respectively, which were high. However, the differences between diets and between traits were not statistically significant (P > 0.05). The heritability estimates for feed efficiency ratio of shrimp fed zero- and high fishmeal diets were 0.186 ±0.098 and 0.341±0.110, respectively, which were low to medium. The differences in feed efficiency ratio between diets was not statistically significant (P > 0.05). All the above heritability estimates were overestimated, to some extent, because common environmental effects were not estimated. Therefore, there was not much genetic variation in feed efficiency ratio in shrimp fed a zero-fishmeal diet, suggesting limited potential in selection breeding of this trait.Regardless of diet, genetic correlations among feed intake, weight gain, and feed efficiency ratio were moderate to high (0.580±0.188 to 0.945±0.040). Genetic correlations between diets were 0.580± 0.171 and 0.676±0.155 for feed intake and weight gain, respectively, indicating that these two traits had moderate genotype by diet interactions. The genetic correlation between diets was only 0.299±0.304 for feed efficiency ratio, which indicates a serious genotype by diet interaction. According to the above results, use of a zero-fishmeal diet is not good for the culture and breeding of P. vannamei. In the future, fishmeal content in the diet may be appropriately increased, and genetic parameters and potential feed efficiency traits should be evaluated in low fishmeal diets.

Litopenaeus vannamei, also known as the Pacific white shrimp or white foot shrimp, is a member of the Crustacea, Decapoda, Palaemonidae, and Litopenaeus families. This species is mainly distributed on the Mexican and Pacific coasts of South America. Owing to its excellent immune characteristics, L. vannamei is currently one of the most important economic shrimp species in China.In the aquaculture process and under natural environmental conditions, low dissolved oxygen (DO) condition or even hypoxia frequently occurs. Under high-density culture operations, acute hypoxia is one of the major factors affecting the survival rate of shrimp and the quality of aquaculture water, causing not only a large number of shrimp deaths in the short term but also water decay and fermentation, increased turbidity, and other phenomena. The genetic improvement of L. vannamei, as well as the screening and breeding of exceptionally tolerant species under low DO conditions, are crucial to the sustainable development of the shrimp industry. Global selective breeding efforts based on quantitative genetics to improve genetic progress in L. vannamei have been extensive, and the main targeted traits were growth, survival, disease resistance, and stress tolerance. Few studies have been conducted on the evaluation of genetic parameters and screening of strains for hypoxic tolerance traits in L. vannamei; in particular, the genetic parameters and cytological characteristics of key tissues of L. vannamei under short-term highly lethal DO levels (0.3–0.5 mg/L) have not been reported.In this study, we used two strains of L. vannamei, GK (a strain with high disease resistance) and K (a strain with fast growth characteristics), each with a total of 6 560 shrimp from 41 families, to count survival traits at highly lethal DO levels, analyze differences in hypoxia tolerance traits between strains and within families of the same strain, and evaluate genetic parameters. Individual gill, muscle, and hepatopancreas tissues from families with significantly different tolerance levels were histologically investigated and compared at the cellular level. The results showed that there was a significant difference (P < 0.05) in semi-lethal survival (SS50) between families within both the GK and K strains when overall semi-lethality was reached under hypoxic environmental conditions of 0.3 mg/L, with SS50 values of 49.30% for GK and 42.52% for K. The coefficients of variation for survival times of families within each strain were 60% and 45% for GK and K, respectively. Using the threshold trait animal model, the genetic parameters were estimated to be between 0.345±0.031 and 0.378±0.029 using the survival status of the individual at semi-lethal levels as the observed value (1 for survival and 0 for death), and the heritability was between 0.219±0.031 and 0.237±0.029 following transformation on a continuously varying scale of observed values, indicating a moderate level of heritability. Moreover, the gill, muscle, and hepatopancreas tissues of the hypoxia-tolerant and hypoxia-sensitive families in the GK strain exhibited varying degrees of damage and different degrees of variations in the physiological characteristics of resistance among different families. This may be related to the resistance tolerance strength of different GK families. In contrast to the gill, muscle, and hepatopancreas tissues of the control shrimp, the equivalent tissues of shrimp from tolerant and sensitive families showed different degrees of changes following stress. The gill lumen of the filaments of the experimental group from sensitive families increased, the number of blood cells increased, the structure of the epithelial layer was gradually destroyed or even disintegrated, the vacuolation of the hepatopancreas was severe, the lumen of the ducts became irregularly deformed, and the muscle tissue muscle bundle interval widened with a certain degree of deformation. These results suggested that there was abundant genetic variation in the tolerance traits of L. vannamei at highly lethal DO levels and that this species was amendable to selective breeding practices. This study provides a reference and basis for the selection and breeding of hypoxia-tolerant traits in L. vannamei.

Plant Elicitor Peptides (Peps) induce plant immune responses and inhibit root growth through their receptors PEPR1 and PEPR2, two receptor-like kinases. In our study, we found a previously unknown function of Peps that enhance root hair growth in a PEPRs-independent manner. When we characterized the expression patterns of PROPEP genes, we found several gene promoters of PROPEP gene family were particularly active in root hairs. Furthermore, we observed that PROPEP2 is vital for root hair development, as disruption of PROPEP2 gene led to a significant reduction in root hair density and length. We also discovered that PROPEP2 regulates root hair formation via the modulation of CPC and GL2 expression, thereby influencing the cell-fate determination of root hairs. Additionally, calcium signaling appeared to be involved in PROPEP2/Pep2-induced root hair growth. These findings shed light on the function of Peps in root hair development.

The current study aimed to provide a precise assessment of the genetic parameters associated with growth and white spot syndrome virus (WSSV) resistance traits in Pacific white shrimp (Litopenaeus vannamei). This was achieved through a controlled WSSV challenge assay and the analysis of phenotypic values of five traits: body weight (BW), overall length (OL), body length (BL), tail length (TL), and survival hour post-infection (HPI). The analysis included test data from a total of 1017 individuals belonging to 20 families, of which 293 individuals underwent whole-genome resequencing, resulting in 18,137,179 high-quality SNP loci being obtained. Three methods, including pedigree-based best linear unbiased prediction (pBLUP), genomic best linear unbiased prediction (GBLUP), and single-step genomic BLUP (ssGBLUP) were utilized. Compared to the pBLUP model, the heritability of growth-related traits obtained from GBLUP and ssGBLUP was lower, whereas the heritability of WSSV resistance was higher. Both the GBLUP and ssGBLUP models significantly enhanced prediction accuracy. Specifically, the GBLUP model improved the prediction accuracy of BW, OL, BL, TL, and HPI by 4.77%, 21.93%, 19.73%, 19.34%, and 63.44%, respectively. Similarly, the ssGBLUP model improved prediction accuracy by 10.07%, 25.44%, 25.72%, 19.34%, and 122.58%, respectively. The WSSV resistance trait demonstrated the most substantial enhancement using both genomic prediction models, followed by body size traits (e.g., OL, BL, and TL), with BW showing the least improvement. Furthermore, the choice of models minimally impacted the assessment of genetic and phenotypic correlations. Genetic correlations among growth traits ranged from 0.767 to 0.999 across models, indicating high levels of positive correlations. Genetic correlations between growth and WSSV resistance traits ranged from (−0.198) to (−0.019), indicating low levels of negative correlations. This study assured significant advantages of the GBLUP and ssGBLUP models over the pBLUP model in the genetic parameter estimation of growth and WSSV resistance in L. vannamei, providing a foundation for further breeding programs.

The Pacific white shrimp, Penaeus vannamei, is highly susceptible to white spot syndrome virus (WSSV). Our study explored the transcriptomic responses of P. vannamei from resistant and susceptible families, uncovering distinct expression patterns after WSSV infection. The analysis revealed a higher number of differentially expressed genes (DEGs) in the susceptible family following WSSV infection compared to the resistant family, when both were evaluated against their respective control groups, indicating that the host resistance of the family line influences the transcriptome. The results also showed that subsequent to an identical duration following WSSV infection, there were more DEGs in P. vannamei with a high viral load than in those with a low viral load. To identify common transcriptomic responses, we profiled DEGs across families at 96 and 228 h post-infection (hpi). The analysis yielded 64 up-regulated and 37 down-regulated DEGs at 96 hpi, with 33 up-regulated and 34 down-regulated DEGs at 228 hpi, showcasing the dynamics of the transcriptomic response over time. Real-time RT-PCR assays confirmed significant DEG expression changes post-infection. Our results offer new insights into shrimp’s molecular defense mechanisms against WSSV.

White spot disease (WSD) outbreaks pose a significant threat to the Pacific white shrimp (Litopenaeus vannamei) farming industry. The causative agent is the white spot syndrome virus (WSSV). There are no effective treatments for WSD so far. Therefore, understanding the resistance mechanisms of L. vannamei against the WSSV is crucial. C-type lectins (CTLs) are important pattern recognition receptors (PRRs) that promote agglutination, phagocytosis, encapsulation, bacteriostasis, and antiviral infections. This study cloned the C-type lectin domain family 4 member F (LvCLEC4F) from L. vannamei. LvCLEC4F contains a 492 bp open reading frame (ORF) encoding a protein of 163 amino acids, including a carbohydrate recognition domain (CRD). Following a challenge with the WSSV, the expression profile of LvCLEC4F was significantly altered. Using RNA interference (RNAi) technology, it was found that LvCLEC4F promotes WSSV replication and affects the expression levels of genes related to the regulation of apoptosis, signaling and cellular stress response, and immune defense. Meanwhile, the hemolymph agglutination phenomenon in vivo was weakened when LvCLEC4F was knocked down. These results indicated that LvCLEC4F may play an important role in the interaction between L. vannamei and WSSV.

Marine animals possess genomes of considerable complexity and heterozygosity. Their unique reproductive system, characterized by high fecundity and substantial early mortality rates, increases the risk of inbreeding, potentially leading to severe inbreeding depression during various larval developmental stages. In this study, we established a set of inbred families of Fenneropenaeus chinensis, with an inbreeding coefficient of 0.25, and investigated elimination patterns and the manifestations of inbreeding depression during major larval developmental stages. Reduced-representation genome sequencing was utilized to explore the genotype frequency characteristics across two typical elimination stages. The results revealed notable mortality in hatching and metamorphosis into mysis and post-larvae stages. Inbreeding depression was also evident during these developmental stages, with depression rates of 24.36%, 29.23%, and 45.28%. Segregation analysis of SNPs indicated an important role of gametic selection before hatching, accounting for 45.95% of deviation in the zoea stage. During the zygotic selection phase of larval development, homozygote deficiency and heterozygote excess were the main selection types. Summation of the two types explained 82.31% and 89.91% of zygotic selection in the mysis and post-larvae stage, respectively. The overall distortion ratio decreased from 22.37% to 12.86% in the late developmental stage. A total of 783 loci were identified through selective sweep analysis. We also found the types of distortion at the same locus could change after the post-larvae stage. The predominant shifts included a transition of gametic selection toward normal segregation and other forms of distortion to heterozygous excess. This may be attributed to high-intensity selection on deleterious alleles and genetic hitchhiking effects. Following larval elimination, a greater proportion of heterozygous individuals were preserved. We detected an increase in genetic diversity parameters such as expected heterozygosity, observed heterozygosity, and polymorphic information content in the post-larvae stage. These findings suggest the presence of numerous recessive deleterious alleles and their linkage and suggest a major role of the partial dominance hypothesis. The results provide valuable insights into the mechanisms of inbreeding depression in marine animals and offer guidance for formulating breeding strategies in shrimp populations.

Temperature is an important environmental factor that affects the growth, development, and metabolism of shrimp. High temperatures tend to cause a decrease in the immunity and production performance of shrimp, resulting in a significant increase in shrimp disease and mortality. Based on transcriptome sequencing data, we screened for the involvement of DNA damage-inducible transcript 4 like (ddit4l) in the temperature stress process of Litopenaeus vannamei. Ddit4l (also known as Redd2 and Rtp801L) is involved in regulating cell survival, proliferation, and apoptosis by affecting DNA damage repair factors, hypoxia-inducible factors, and other signaling pathways when stimulated by DNA damage, hypoxia, ischemia, and oxidative stress, among others. Ddit4l may be an important transducer of pathological stress in autophagy through mammalian target of rapamycin (mTOR) signaling. Autophagy is a process in which cells use lysosomes to degrade their own damaged organelles and macromolecules, and the process plays an important role in maintaining the balance of protein metabolism and the stability of the intracellular environment, promoting cell growth and development. An important regulator of autophagy is the mTOR, which responds to various stimuli, such as growth factors, cellular energy status, oxygen concentrations, and stress, to regulate cell metabolism and growth. Ddit4l and its homolog ddit4 are upstream mTOR inhibitors in several tissues and cell models. We set up sudden temperature change and gradual temperature change experiments to induce a stress response in L. vannamei. In the gradual change experiment, the water temperature was increased from 26℃ to 38℃ at a rate of 4℃ per day, and cooled back to 26℃ at the same rate. Two individuals were randomly selected from each of the three parallel experiments in the experimental group at each time point. The eyestalk, gill, hepatopancreas, nerve, and muscle were used for subsequent RNA extraction and gene expression quantification. RNA quality and concentration were also examined. The partial ddit4l cDNA (complete ORF and partial UTR) of L. vannamei was obtained directly by PCR amplification to evaluate the structure and function of the Lv-ddit4l gene. The Lv-ddit4l sequence contained a 495 bp open reading frame, encoding 164 amino acids, with a molecular weight of 18.51 kDa, and an isoelectric point of 5.35. The instability index was 65.83, which classifies the protein as unstable. Lv-ddit4l contains a highly conserved RTP801-C domain. BLAST alignment and phylogenetic tree analysis showed that the Lv-ddit4l gene had 59.76% similarity with the Chionoecetes opilio ddit4l gene, and first clustered with C. opilio. A real-time RT-PCR confirmatory experiment considered three parallel groups for each sample with 18S rRNA as the reference gene. Relative gene expression was analyzed using the 2–ΔΔCt method. Quantitative analysis of gene expression levels showed that Lv-ddit4l was widely expressed in all of the examined tissues, with similar expression levels at 26℃. When the temperature was increased gradually from 26℃ to 32℃, the Lv-ddit4l gene in the eyestalk was significantly increased (P < 0.05), and was 10.37 times higher than that in the control (A-26℃). Therefore, this gene is hypothesized to respond to temperature changes in the eyestalk of L. vannamei. The expression of Lv-ddit4l in the eyestalk, gills, hepatopancreas, nerves, and muscles did not change significantly when the water temperature was gradually increased to the highest temperature of 38℃; it is speculated that this gene may play a role in pre-heat stress. When the temperature was gradually cooled from 38℃ to 32℃, the expression of Lv-ddit4l increased significantly in the gill and hepatopancreas (P < 0.05), which were respectively 3.37 and 2.03 times higher than that of the control group (A-26℃). There was no significant difference in gene expression under different levels of gradual changes in temperature stress in the nerve and muscle. The expression in the gill, hepatopancreas, and nerve was significantly up-regulated (P < 0.05) at a sudden change in water temperature to 36℃ for 10 min, and the change in expression in the hepatopancreas was 24.06 times higher than that in the control group (A-26℃). Gene expression in the gills and nerves was 1.22-fold and 3.67-fold higher than that in the control (A-26℃), respectively. When the water temperature was increased to 36℃ suddenly for 24 h, Lv-ddit4l gene expression increased significantly in the muscle (P < 0.05). Lv-ddit4l showed significant up-regulation of expression in the gill, hepatopancreas, nerve, and muscle with sudden changes in temperature, suggesting that this gene may play a role in the stress induced by sudden increases in temperature. When the temperature was restored from 38℃ and 36℃ to 26℃, the expression of the Lv-ddit4l gene was downregulated in L. vannamei. The above results indicate that Lv-ddit4l is associated with the response to high-temperature stress.

Potassium is essential for plant growth and development. Here the authors present evidence that plants respond to low potassium availability by modulating the abundance and phosphorylation status of proteins in CBL-CIPK-channel modules.

The Pacific whiteleg shrimp (Penaeus vannamei) is a highly significant species in shrimp aquaculture. In the production of shrimp larvae, noticeable variations in the reproductive capacity among female individuals have been observed. Some females experience slow gonadal development, resulting in the inability to spawn, while others undergo multiple maturations and contribute to the majority of larval supply. Despite numerous studies that have been conducted on the regulatory mechanisms of ovarian development in shrimp, the factors contributing to the differences in reproductive capacity among females remain unclear. To elucidate the underlying mechanisms, this study examined the differences in the ovarian characteristics between high and low reproductive bulks at different maturity stages, focusing on the cellular and molecular levels. Transmission electron microscopy analysis revealed that the abundance of the endoplasmic reticulum, ribosomes, mitochondria, and mitochondrial cristae in oocytes of high reproductive bulk was significantly higher than that of the low reproductive bulk in the early stages of ovarian maturation (stages I and II). As the ovaries progressed to late-stage maturation (stages III and IV), differences in the internal structures of oocytes between females with different reproductive capacities gradually diminished. Transcriptome analysis identified differentially expressed genes (DEGs) related to the mitochondria between two groups, suggesting that energy production processes might play a crucial role in the observed variations in ovary development. The expression levels of the ETS homology factor (EHF) and PRDI-BF1 and RIZ homology domain containing 9 (PRDM9), which were significantly different between the two groups, were compared using qRT-PCR in individuals at different stages of ovarian maturation. The results showed a significantly higher expression of the EHF gene in the ovaries of high reproductive bulk at the II and IV maturity stages compared to the low reproductive bulk, while almost no expression was detected in the eyestalk tissue of the high reproductive bulk. The PRDM9 gene was exclusively expressed in ovarian tissue, with significantly higher expression in the ovaries of the high reproductive bulk at the four maturity stages compared to the low reproductive bulk. Fluorescence in situ hybridization further compared the expression patterns of EHF and PRDM9 in the ovaries of individuals with different fertility levels, with both genes showing stronger positive signals in the high reproductive bulk at the four ovarian stages. These findings not only contribute to our understanding of the regulatory mechanisms involved in shrimp ovarian development, but also provide valuable insights for the cultivation of new varieties aimed at improving shrimp fecundity.

To explore the performance differences among offspring obtained by the intra- and inter-strain crosses, strain A with outstanding WSSV resistance and strain B with outstanding growth of Litopenaeus vannamei were used to construct four experimental groups in a complete diallel cross. 27 growth-related phenotypic traits and WSSV resistance represented by the surviving hours post-infection were obtained via the growth and the virus challenge test. Mixed linear model along with physical pedigree were utilized to calculate the least squares means, correlations, heritability and heterosis. The results showed that crossbreeding can realize character complementation of advantage strain. The performance of WSSV resistance was ranked AA> BA> AB> BB, and the reciprocal crosses performed significantly better than intra-strain group BB. Most growth-related traits in two reciprocal cross groups were also significantly better than those in AA group. Comparisons between the two reciprocal crosses showed that BA cross was better than AB in terms of WSSV resistance and growth. Heterosis of WSSV resistance in BA cross was 5.74∼10.18% greater than that in AB, meanwhile the absolute weight gain rate of BA cross was 4.83% higher than that of AB. The heritability estimates of body length and body weight were 0.0278 and 0.339, both of which reached medium level. The genetic and phenotypic correlations among growth-related traits were from 0.36 to 0.99 and 0.32 to 0.99, respectively. In term of WSSV resistance, the heritability estimate was 0.135 ± 0.082. The genetic and phenotypic correlations between WSSV resistance and growth-related traits were from − 0.034 to − 0.573 and − 0.001 to − 0.202, respectively. The results showed that hybrid offspring obtained by crossing female parents with outstanding WSSV resistance and male parents with outstanding growth culminated in a superior heterosis in both traits. This study provides basic reference data for L. vannamei breeding of new strains that are disease resistant and fast-growing.

The relationship between inbreeding and traits of disease resistance in shrimp has not been studied comprehensively. To explore the effect of inbreeding on white spot syndrome virus (WSSV) resistance in Fenneropenaeus chinensis, we established F. chinensis “Huanghai No. 5” populations with different inbreeding coefficients. We infected F. chinensis individuals with WSSV to compare WSSV-resistance between five inbred groups and a control group. Survival time, survival rate, distribution of deaths, and cumulative mortality rate were analyzed. A mixed linear model was established based on the acquired data. The viral load in muscle tissue of F. chinensis of 12 families from the five inbred groups and the control group was determined at different time points via real-time PCR. The average survival and longest survival time of the inbred groups were lower than those of the control group, and the coefficient of variation was universally greater than that of the control group. The least squares means of survival time of the five inbred groups (F0.1325, F0.25, F0.3125, F0.375, F0.5) were 118.96 115.14, 124.58, 123.97, and 101.34 h, respectively, which were 10.28%, 13.16%, 6.04%, 6.5%, and 23.57% lower than that of the control group. Further, when the inbreeding coefficient increased by 10%, WSSV infection survival time was changed by − 7.76%, − 5.26%, − 1.93%, − 1.73%, and − 4.71% for the five inbred groups, respectively. Thus, the results indicated that inbreeding enhanced the susceptibility of F. chinensis to WSSV infection, and the resistance of F. chinensis to WSSV declined as the inbreeding coefficient values increased, consequently, resulting in greater population dispersion, intensified differentiation, and instability of resistant phenotypes. In conclusion, we elucidated the effect of inbreeding on F. chinensis resistance to WSSV infection and established a theoretical basis to conduct further research in the future.

Penaeusvannamei is the most important economic shrimp in the world. Many selective breeding programs are carried out to improve its production and performance traits. Although significant differences in the reproductive ability of female P. vannamei under artificial breeding conditions have been reported, the genome-wide adaption of the reproductive ability of domesticated female P. vannamei is less investigated. In this study, whole-genome analysis was performed along with pooled DNA sequencing on two fecundity separated bulks, high fecundity bulk (HB), and low fecundity bulk (LB). Each bulk contained 30 individuals from 3 commercial populations. A sequencing depth of >30× was achieved for each bulk, leading to the identification of 625,181 and 629,748 single nucleotide polymorphisms (SNPs) in HB and LB, respectively. Fixation index (Fst) combined with p ratio allowed for the identification of 145 selective sweep regions, with a sequence length of 14.5 Mb, accounting for 0.59% of the genome. Among the 145 selective sweep regions, a total of 64,046 SNPs were identified, and further verification was performed by genotyping 50 candidate SNPs on 60 samples from the offspring of the three populations. Furthermore, 121 genes were screened from the sweep regions. GO annotation and KEGG enrichment analyses showed that partial genes were essential for fecundity regulation. This study provides important information for in-depth investigation of genomic characteristics for long-term selective breeding on the fecundity of female P. vannamei and will also be important for genome-assisted breeding of P. vannamei in the future.

Abstract Fenneropenaeus chinensis is one of the most important aquaculture species in China. Research on its genomic and genetic structure not only helps us comprehend the genetic basis of complex economic traits, but also offers theoretical guidance in selective breeding. In the present study, a genome survey sequencing was performed to generate a rough reference genome utilized for groping preliminary genome characteristics and facilitate linkage and quantitative trait locus (QTL) mapping. Linkage mapping was conducted using a reduced-representation sequencing method 2b-RAD. In total, 36,762 SNPs were genotyped from 273 progenies in a mapping family, and a high-resolution linkage map was constructed. The consensus map contained 12,884 markers and spanned 5257.81 cM with an average marker interval of 0.41 cM, which was the first high-resolution genetic map in F. chinensis to our knowledge. QTL mapping and association analysis were carried out in 29 characters including body size, sex and disease resistance. 87 significant QTLs were detected in several traits and they were also evaluated by association analysis. Results of this study provide us valuable suggestions in genetic improvement and breeding of new varieties and also lay a basic foundation for further application of cloning of economic genes in selective breeding program and marker-assisted selection.

Abstract Background Photosystem II (PSII) is a highly conserved integral-membrane multi-subunit pigment-protein complex. The proteins, pigments, lipids, and ions in PSII need to be assembled precisely to ensure a proper PSII biogenesis. D1 is the main subunit of PSII core reaction center (RC), and is usually synthesized as a precursor D1. D1 maturation by the C-terminal processing protease CtpA is essential for PSII assembly. However, the detailed mechanism about how D1 maturation affects PSII assembly is not clearly elucidated so far. In this study, Arabidopsis thaliana CtpA mutant (atctpa: SALK_056011), which lacks the D1 mature process, was used to investigate the function of this process on PSII assembly in more details. Results Without the C-terminal processing of precursor D1, PSII assembly, including PSII monomer, dimer, especially PSII supercomplexes (PSII SCs), was largely compromised as reported previously. Western blotting following the BN-2D-SDS PAGE revealed that although the assembly of PSII core proteins D2, CP43 and CP47 was affected by the loss of D1 mature process, the incorporation of CP43 was affected the most, indicated by its most reduced assembly efficiency into PSII SCs. Furthermore, the slower growth of yeast cells which were co-transformed with pD1 and CP43, when compared with the ones co-transformed with mature D1 and CP43, approved the existence of D1 C-terminal tail hindered the interaction efficiency between D1 and CP43, indicating the physiological importance of D1 mature process on the PSII assembly and the healthy growth of the organisms. Conclusions The knockout Arabidopsis atctpa mutant is a good material to study the unexpected link between D1 maturation and PSII SCs assembly. The loss of D1 maturation mainly affects the incorporation of PSII core protein CP43, an inner antenna binding protein, which functions in the association of LHCII complexes to PSII dimers during the formation of PSII SCs. Our findings here provide detailed supports of the role of D1 maturation during PSII SCs assembly in higher plants.

Abstract Segregation distortion is a common phenomenon found in most genetic mapping studies and is an important resource to dissect the mechanism of action in gene loci that cause deviation. Marine animals possess high genetic diversity and genomic heterozygosity, they therefore are ideal model organisms to study segregation distortion induced by selection. In the present study, we constructed a full-sib family of Fenneropenaeus chinensis and exerted high-intensity selection on 10,000 incipient progenies. 2b-RAD method was employed in remaining 273 individuals to develop genome-wide SNPs for segregating analysis and 41,612 SNPs were developed. 50.77% of 32,229 high-quality representative markers deviated from the expected Mendelian ratio. Results showed that most of these distorted markers (91.57%) were influenced at zygotic level. Heterozygote excess (53.07%) and homozygous deletions (41.96%) may both play an important role, sum of which explained 95.03% of distortion after fertilization. However, further results identified highly probable linkage among deleterious alleles, which may account for a considerable portion of heterozygote excess rather than single locus with heterozygote advantage. Results of this study support a major role of deleterious alleles in genetic load, thus in favor of partial dominance hypothesis. It would also offer necessary recommendations for the formulation of breeding strategy in shrimps.

White spot syndrome virus (WSSV) is one of the most lethal viruses in shrimp aquaculture. Compared with other shrimp species, the Chinese shrimp (Fenneropenaeus chinensis) is more susceptible to WSSV. The mortality of WSSV-infected F. chinensis can reach 100% within a week. No effective solution currently exists to solve the WSSV problem in F. chinensis aquaculture. After years of research, we found that individual F. chinensis from the same family can carry different viral load at the same time post-WSSV infection. To better understand the anti-WSSV mechanism of F. chinensis, we analysed the transcriptomic data from F. chinensis with high and low WSSV load at the same time post-WSSV infection and from healthy F. chinensis, in a WSSV-resistant group and a WSSV-susceptible group, respectively. We identified more differentially expressed genes in F. chinensis with high WSSV load than in those with low WSSV load at the same time post-WSSV infection. Many differentially expressed genes were enriched in Gene Ontology terms related to binding function and the extracellular matrix-receptor interaction pathway, which might play important roles in the anti-WSSV processes among F. chinensis.

Summary: Symbioses between angiosperms and rhizobia or arbuscular mycorrhizal fungi are controlled through a conserved signaling pathway. Microbe-derived, chitin-based elicitors activate plant cell surface receptors and trigger nuclear calcium oscillations, which are decoded by a calcium/calmodulin-dependent protein kinase (CCaMK) and its target transcription factor interacting protein of DMI3 (IPD3). Genes encoding CCaMK and IPD3 have been lost in multiple non-mycorrhizal plant lineages yet retained among non-mycorrhizal mosses. Here, we demonstrated that the moss Physcomitrium is equipped with a bona fide CCaMK that can functionally complement a Medicago loss-of-function mutant. Conservation of regulatory phosphosites allowed us to generate predicted hyperactive forms of Physcomitrium CCaMK and IPD3. Overexpression of synthetically activated CCaMK or IPD3 in Physcomitrium led to abscisic acid (ABA) accumulation and ectopic development of brood cells, which are asexual propagules that facilitate escape from local abiotic stresses. We therefore propose a functional role for Physcomitrium CCaMK-IPD3 in stress-associated developmental reprogramming

Abstract Background Competition is a common social interaction among shrimp and depending on its intensity, it can affect heritable variation and response to selection. Little is known about the variance of indirect genetic effects (IGE) under competitive and non-competitive conditions in shrimp. In this study, we used extended mixed linear models to estimate genetic parameters for the direct genetic effect (DGE) and IGE on body weight in Litopenaeus vannamei raised under ad libitum (AF, non-competitive environment) and restricted (RF, competitive environment) feeding regimes. Results Estimates of heritabilities for body weight obtained with a traditional animal model (i.e. without accounting for IGE) were 0.11 ± 0.09 under AF and 0.25 ± 0.11 under RF. With extended animal models that accounted for IGE, the corresponding estimates for body weight were 0.07 ± 0.08 and 0.34 ± 0.11. Thus, heritabilities were higher under the RF regime than under the AF regime, regardless of whether IGE was accounted for or not. The log-likelihood ratio test revealed significant IGE under the RF regime. Although estimates of indirect genetic variance were low (0.0023 ± 0.0013 for AF and 0.0028 ± 0.0012 for RF), they contributed substantially to the total heritable variance: 66.8% for AF and 692.2% for RF. The total heritable variance was smaller under the RF regime (0.7 ± 1.3) than under the AF regime (5.8 ± 2.6) because of the high contribution of the negative covariance between DGE and IGE (− 7.03). Estimates of the correlation between DGE and IGE were 0.32 ± 0.47 under AF and − 0.93 ± 0.15 under RF, those of DGE and IGE for body weight between both regimes were 0.94 ± 0.07 and 0.67 ± 0.20, respectively, and those of IGE for body weight with DGE for survival were − 0.12 ± 0.22 under AF and − 0.58 ± 0.20 under RF. Conclusions These results indicate that strong competitive interactions occurred under the RF regime in L. vannamei. Significant reranking and variation in IGE of individuals were observed between the two feeding regimes. Strong competitive interactions reduced the total heritable variation for body weight when food was restricted. These results indicate that the extent of competition among L. vannamei depends on the feeding regime applied and that this competition affects the genetic basis of body weight.

Abstract Background The receptor-like kinase FEROINA (FER) plays a crucial role in controlling plant vegetative growth partially by sensing the rapid alkalinization factor (RALF) peptide. However, the role of RALF1-FER in the vegetative-reproductive growth transition remains unknown. Here, we analyze the mechanism through which FER affects the flowering time in Arabidopsis. Results We found that the FER mRNA levels exhibit an oscillating pattern with a diurnal rhythm and that the clock oscillator CIRCADIAN CLOCK-ASSOCIATED1 (CCA1) up-regulates the expression of FER by associating with its chromatin. In addition, FER expression is regulated by clock genes, and FER also modulates the expression patterns of clock genes. Consistent with its gene expression pattern, FER positively regulates flowering by modulating the transcript accumulation and mRNA alternative splicing of certain flowering-related genes, including FLOWERING LOCUS C (FLC) and its homolog MADS AFFECTING FLOWERING (MAF). However, the RALF1 ligand negatively regulates flowering compared with FER. Conclusions We found that FER, which is up-regulated by CCA1, controls the flowering time by regulating the transcript accumulation and mRNA alternative splicing (AS) of some important flowering genes, and these findings link FER to the floral transition.

Evolution of adaptive mechanisms to abiotic stress is essential for plant growth and development. Plants adapt to stress conditions by activating the abscisic acid (ABA) signaling pathway. It has been suggested that the ABA receptor, clade A protein phosphatase, SnRK2 type kinase, and SLAC1 anion channel are important components of the ABA signaling pathway. In this study, we report that the shaker type potassium (K+) channel, GORK, modulates plant responses to ABA and abiotic stresses. Our results indicate that the full length of PP2CA is needed to interact with the GORK C-terminal region. We identified a loss of function allele in gork that displayed ABA-hyposensitive phenotype. gork and pp2ca mutants showed opposite responses to ABA in seed germination and seedling growth. Additionally, gork mutant was tolerant to the NaCl and mannitol treatments, whereas pp2ca mutant was sensitive to the NaCl and mannitol treatments. Thus, our results indicate that GORK enhances the sensitivity to ABA and negatively regulates the mechanisms involved in high salinity and osmotic stresses via PP2CA-mediated signals.

Soil salinity has become a major stress factor that reduces crop productivity worldwide. Sodium (Na+) toxicity in a number of crop plants is tightly linked with shoot Na+ overaccumulation, thus Na+ exclusion from shoot is crucial for salt tolerance in crops. In this study, we identified a member of the high-affinity K+ transport family (HAK), OsHAK12, which mediates shoots Na+ exclusion in response to salt stress in rice. The Oshak12 mutants showed sensitivity to salt toxicity and accumulated more Na+ in the xylem sap, leading to excessive Na+ in the shoots and less Na+ in the roots. Unlike typical HAK family transporters that transport K+, OsHAK12 is a Na+-permeable plasma membrane transporter. In addition, OsHAK12 was strongly expressed in the root vascular tissues and induced by salt stress. These findings indicate that OsHAK12 mediates Na+ exclusion from shoot, possibly by retrieving Na+ from xylem vessel thereby reducing Na+ content in the shoots. These findings provide a unique function of a rice HAK family member and provide a potential target gene for improving salt tolerance of rice.

Potassium (K+) levels in the soil often limit plant growth and development. As a result, crop production largely relies on the heavy use of chemical fertilizers, presenting a challenging problem in sustainable agriculture. To breed crops with higher K+-use efficiency (KUE), we must learn how K+ is acquired from the soil by the root system and transported to the rest of the plant through K+ transporters. In this study, we identified the function of the rice K+ transporter OsHAK8, whose expression level is downregulated in response to low-K+ stress. When OsHAK8 was disrupted by CRISPR/Cas9-mediated mutagenesis, Oshak8 mutant plants showed stunted growth, especially under low-K+ conditions. Ion content analyses indicated that K+ uptake and root-to-shoot K+ transport were significantly impaired in Oshak8 mutants under low-K+ conditions. As the OsHAK8 gene was broadly expressed in different cell types in the roots and its protein was targeted to the plasma membrane, we propose that OsHAK8 serves as a major transporter for both uptake and root-to-shoot translocation in rice plants.

Nitrogen (N), phosphorus (P), and potassium (K) are three major macronutrients essential for plant life. These nutrients are acquired and transported by several large families of transporters expressed in plant roots. However, it remains largely unknown how these transporters are distributed in different cell-types that work together to transfer the nutrients from the soil to different layers of root cells and eventually reach vasculature for massive flow. Using the single cell transcriptomics data from Arabidopsis roots, we profiled the transcriptional patterns of putative nutrient transporters in different root cell-types. Such analyses identified a number of uncharacterized NPK transporters expressed in the root epidermis to mediate NPK uptake and distribution to the adjacent cells. Some transport genes showed cortex- and endodermis-specific expression to direct the nutrient flow toward the vasculature. For long-distance transport, a variety of transporters were shown to express and potentially function in the xylem and phloem. In the context of subcellular distribution of mineral nutrients, the NPK transporters at subcellular compartments were often found to show ubiquitous expression patterns, which suggests function in house-keeping processes. Overall, these single cell transcriptomic analyses provide working models of nutrient transport from the epidermis across the cortex to the vasculature, which can be further tested experimentally in the future.

Mitochondrial transcription termination factors (mTERFs) are highly conserved proteins in metazoans. Plants have many more mTERF proteins than animals. The functions and the underlying mechanisms of plants’ mTERFs remain largely unknown. In plants, mTERF family proteins are present in both mitochondria and plastids and are involved in gene expression in these organelles through different mechanisms. In this study, we screened Arabidopsis mutants with pigment-defective phenotypes and isolated a T-DNA insertion mutant exhibiting seedling-lethal and albino phenotypes [seedling lethal 1 (sl1)]. The SL1 gene encodes an mTERF protein localized in the chloroplast stroma. The sl1 mutant showed severe defects in chloroplast development, photosystem assembly, and the accumulation of photosynthetic proteins. Furthermore, the transcript levels of some plastid-encoded proteins were significantly reduced in the mutant, suggesting that SL1/mTERF3 may function in the chloroplast gene expression. Indeed, SL1/mTERF3 interacted with PAP12/PTAC7, PAP5/PTAC12, and PAP7/PTAC14 in the subgroup of DNA/RNA metabolism in the plastid-encoded RNA polymerase (PEP) complex. Taken together, the characterization of the plant chloroplast mTERF protein, SL1/mTERF3, that associated with PEP complex proteins provided new insights into RNA transcription in the chloroplast.

Using pooled DNA genotyping to estimate the proportional contributions from multiple families in a pooled sample is of particular interest for selective breeding in aquaculture. We compared different pooled libraries with separate 2b-RAD sequencing of Litopenaeus vannamei individuals to assess the effect of different population structures (different numbers of individuals and families) on pooled DNA sequencing, the accuracy of parent sequencing of the DNA pools and the effect of SNP numbers on pooled DNA sequencing. We demonstrated that small pooled DNA genotyping of up to 53 individuals by 2b-RAD sequencing could provide a highly accurate assessment of population allele frequencies. The accuracy increased as the number of individuals and families increased. The allele frequencies of the parents from each pool were highly correlated with those of the pools or the corresponding individuals in the pool. We chose 500-28,000 SNPs to test the effect of SNP number on the accuracy of pooled sequencing, and no linear relationship was found between them. When the SNP number was fixed, increasing the number of individuals in the mixed pool resulted in higher accuracy of each pooled genotyping. Our data confirmed that pooled DNA genotyping by 2b-RAD sequencing could achieve higher accuracy than that of individual-based genotyping. The results will provide important information for shrimp breeding programs.

Membrane transport processes are indispensable for many aspects of plant physiology including mineral nutrition, solute storage, cell metabolism, cell signaling, osmoregulation, cell growth, and stress responses. Completion of genome sequencing in diverse plant species and the development of multiple genomic tools have marked a new era in understanding plant membrane transport at the mechanistic level. Genes coding for a galaxy of pumps, channels, and carriers that facilitate various membrane transport processes have been identified while multiple approaches are developed to dissect the physiological roles as well as to define the transport capacities of these transport systems. Furthermore, signaling networks dictating the membrane transport processes are established to fully understand the regulatory mechanisms. Here, we review recent research progress in the discovery and characterization of the components in plant membrane transport that take advantage of plant genomic resources and other experimental tools. We also provide our perspectives for future studies in the field.

Abstract Background Residual feed intake (RFI) was investigated as a measure of feed efficiency in a breeding population of Litopenaeus vannamei. Shrimp from 34 families were housed individually and feed efficiency and growth traits were recorded during two successive growth periods. The objectives of this study were (1) to estimate the heritability of RFI and related traits, including feed efficiency ratio (FER), average daily gain (ADG) and daily feed intake (DFI), (2) to determine the relationships between RFI and other traits, and (3) to evaluate the variation of these traits across two growth periods. Results Shrimp displayed large inter-individual variation in RFI, FER, ADG and DFI during each growth period. Heritability estimates of all these traits during both periods reached high values (0.577 ± 0.232 to 0.707 ± 0.252). RFI showed weak and no genetic correlations with ADG during the two growth periods between days 1 to 21 (0.135 ± 0.204) and 22 to 42 (–0.018 ± 0.128), respectively, but high positive genetic correlations with DFI (>0.8). Weak and moderate negative genetic correlations were observed between RFI and FER during the two periods (–0.126 ± 0.208 and –0.387 ± 0.183). As evidenced by the high genetic correlations between the two periods for each trait (>0.6), trait performance of the shrimp tended to be consistent across periods. Conclusions For the first time, accurate measurement of individual feed efficiency on a large scale was achieved in shrimp. Although the estimated heritability reported here for RFI may be overestimated, it is a heritable trait in L. vannamei that can be improved by genetic improvement. For L. vannamei, the biggest potential advantage in using RFI as a measure of feed efficiency is that it is independent of growth rate, and thus genetic selection on RFI has the potential to improve feed efficiency and reduce feed intake, without compromising growth performance.

A multi-trait selective breeding program of Macrobrachium rosenbergii was initiated in China in 2015. In this program, the M. rosenbergii resources were widely collected from four countries, the origin of the founders was verified with 16 microsatellites and the pedigree was reconstructed, and the optimum contribution selection was used to make the mating design. In this study, we evaluated the genetic parameters and selection response for the harvest body weight (HBW) of M. rosenbergii after being communally reared for 95-109 days. The data were collected from two generations that comprised 25,212 progenies from 150 sires and 198 dams. The residual maximum-likelihood methodology was employed to evaluate the variance components, by fitting an animal model. The accuracy of estimated breeding values increased by 0.38% after pedigree reconstruction using microsatellite markers. The estimated heritability (h2) for HBW was moderate (0.212 ± 0.049) and the common environmental coefficient (c2) was low (0.063 ± 0.017) when all the data were used for the analysis. Within generations, h2 was moderate to high (0.198 ± 0.080 to 0.338 ± 0.049). c2 could only be estimated in G1, which was 0.055 ± 0.030. The average HBW of males was significantly larger than that of females (P < 0.01). h2 estimated for female HBWs were higher than that for males within generations, while h2 estimated for female HBWs were lower than that for males across generations. But they were not significantly different (P > 0.05). The genetic correlations between sexes were moderate to high within each generation (0.529 to 0.763). Two methods were used to estimate the realized response. One method was calculated from the differences between the least squares means of the selected population HBW and that of control population HBW, which was 14.01%. The other method was calculated from the differences between the EBVs of the selected population HBW and that of control population HBW, which was 11.52%. The predicted responses derived from two sets of genetic parameters acquired from within- and across- generation datasets were 11.68% and 10.67%, respectively. The present study provides valuable information for breeding programs of M. rosenbergii.

In order to screen the candidate genes of Fenneropenaeus chinensis related to low-temperature tolerance, this research takes juvenile prawns of F. chinensis (P40) in low temperature stress group (4°C) and normal temperature group (18°C) as experimental materials. The results showed that a total of 127,939 Unigenes with average length of 1,190 bp were obtained by assembly, of which 46% were annotated in the Nr database. A total of 1,698 differentially expressed genes were screened by differential gene expression analysis, of which 920 genes showed up-regulated expression and 778 genes showed down-regulated expression. Both GO and KEGG enrichment analysis revealed that differentially expressed genes were enriched in spliceosomes, ribosomes, bile secretion, ABC transport pathways, and cellular nitrogen compound synthesis. A further in-depth analysis obtained 8 genes that may be associated with low-temperature traits of F. chinensis. Five of them displayed up-regulated expression, including ATP-binding cassette protein C, acid ceramidase, glutathione transferase, C-type lectin and heat shock protein HSP70. The remaining three genes, γ-butyl betaine hydroxylase, β-hexosaminidase A and long chain fatty acid-CoA ligase displayed down-regulated expression. Eight differentially expressed genes were randomly selected and the real time RT-PCR verification showed that their expression levels were consistent with the sequencing results, demonstrating the accuracy of the sequencing results. The results of this study provide basic data for revealing the molecular mechanisms of F. chinensis in response to low temperature stress and the molecular assisted breeding of F. chinensis in low temperature.

FERONIA (FER), a plasma membrane receptor-like kinase, is a central regulator of cell growth that integrates environmental and endogenous signals. A peptide ligand rapid alkalinization factor 1 (RALF1) binds to FER and triggers a series of downstream events, including inhibition of Arabidopsis H+-ATPase 2 activity at the cell surface and regulation of gene expression in the nucleus. We report here that, upon RALF1 binding, FER first promotes ErbB3-binding protein 1 (EBP1) mRNA translation and then interacts with and phosphorylates the EBP1 protein, leading to EBP1 accumulation in the nucleus. There, EBP1 associates with the promoters of previously identified RALF1-regulated genes, such as CML38, and regulates gene transcription in response to RALF1 signaling. EBP1 appears to inhibit the RALF1 peptide response, thus forming a transcription-translation feedback loop (TTFL) similar to that found in circadian rhythm control. The plant RALF1-FER-EBP1 axis is reminiscent of animal epidermal growth factor receptor (EGFR) signaling, in which EGF peptide induces EGFR to interact with and phosphorylate EBP1, promoting EBP1 nuclear accumulation to control cell growth. Thus, we suggest that in response to peptide signals, plant FER and animal EGFR use the conserved key regulator EBP1 to control cell growth in the nucleus.

Plants encounter a variety of mechanical stimuli during their growth and development. It is currently believed that mechanosensitive ion channels play an essential role in the initial perception of mechanical force in plants. Over the past decade, the study of Piezo, a mechanosensitive ion channel in animals, has made significant progress. It has been proved that the perception of mechanical force in various physiological processes of animals is indispensable. However, little is still known about the function of its homologs in plants. In this study, by investigating the function of the AtPiezo gene in the model plant Arabidopsis thaliana, we found that AtPiezo plays a role in the perception of mechanical force in plant root cap and the flow of Ca2+ is involved in this process. These findings allow us to understand the function of AtPiezo from the perspective of plants and provide new insights into the mechanism of plant root cap in response to mechanical stimuli.

Phosphate transporters (PHTs) play pivotal roles in phosphate (Pi) acquisition from the soil and distribution throughout a plant. However, there is no comprehensive genomic analysis of the PHT families in Camelina sativa, an emerging oilseed crop. In this study, we identified 73 CsPHT members belonging to the five major PHT families. A whole-genome triplication event was the major driving force for CsPHT expansion, with three homoeologs for each Arabidopsis ortholog. In addition, tandem gene duplications on chromosome 11, 18 and 20 further enlarged the CsPHT1 family beyond the ploidy norm. Phylogenetic analysis showed clustering of the CsPHT1 and CsPHT4 family members into four distinct groups, while CsPHT3s and CsPHT5s were clustered into two distinct groups. Promoter analysis revealed widespread cis-elements for low-P response (P1BS) specifically in CsPHT1s, consistent with their function in Pi acquisition and translocation. In silico RNA-seq analysis revealed more ubiquitous expression of several CsPHT1 genes in various tissues, whereas CsPHT2s and CsPHT4s displayed preferential expression in leaves. While several CsPHT3s were expressed in germinating seeds, most CsPHT5s were expressed in floral and seed organs. Suneson, a popular Camelina variety, displayed better tolerance to low-P than another variety, CS-CROO, which could be attributed to the higher expression of several CsPHT1/3/4/5 family genes in shoots and roots. This study represents the first effort in characterizing CsPHT transporters in Camelina, a promising polyploid oilseed crop that is highly tolerant to abiotic stress and low-nutrient status, and may populate marginal soils for biofuel production.

Plant elicitor peptides (Peps) are damage/danger-associated molecular patterns (DAMPs) that are perceived by a pair of receptor-like kinases, PEPR1 and PEPR2, to enhance innate immunity and induce the growth inhibition of root in Arabidopsis thaliana. In this study, we show that PEPR1 and PEPR2 function vitally in roots to regulate the root immune responses when treating the roots with bacterial pathogen Pst DC3000. PEPR2, rather than PEPR1, played a predominant role in the perception of Pep1 in the roots and further triggered a strong ROS accumulation—the substance acts as an antimicrobial agent or as a secondary messenger in plant cells. Consistently, seedlings mutating two major ROS-generating enzyme genes, respiratory burst oxidase homologs D and F (RBOHD and RBOHF), abolished the root ROS accumulation and impaired the growth inhibition of the roots induced by Pep1. Furthermore, we revealed that botrytis-induced kinase 1 (BIK1) physically interacted with PEPRs and RBOHD/F, respectively, and served downstream of the Pep1-PEPRs signaling pathway to regulate Pep1-induced ROS production and root growth inhibition. In conclusion, this study demonstrates a previously unrecognized signaling crosstalk between Pep1 and ROS signaling to regulate root immune response and root growth.

The Arabidopsis genome comprises eighty genes encoding BTB (broad-complex, tramtrack, and bric-a-brac) family proteins that are characterized with the BTB domain and that potentially serve as substrate adaptors for cullin-based E3-ligases. In addition to the BTB domain, most BTB proteins also contain various other interaction motifs that probably act as target recognition elements. Here, we report three members of the BTB-A2 subfamily that distinctly only contain the BTB domain, BTB-A2.1, BTB-A2.2, and BTB-A2.3, that negatively regulates abscisic acid (ABA) signaling in Arabidopsis. BTB-A2.1, BTB-A2.2, and BTB-A2.3 encoded cytoplasm- and nucleus-localized proteins and displayed highly overlapping expression patterns in Arabidopsis tissues. Disruption of these three genes, but not single or double mutants, resulted in a decrease in ABA-induced inhibition of seed germination. Further analyses demonstrated the expression levels of these three genes were up-regulated by ABA, and their mutation increased ABA signalling. Importantly, protein-protein interaction assays showed that these three BTB-A2 proteins physically interacted with SnRK2.3. Moreover, biochemical and genetic assays indicated that BTB-A2.1, BTB-A2.2, and BTB-A2.3 decreased the stability of SnRK2.3 and attenuated the SnRK2.3 responsible for the ABA hypersensitive phenotype of seed germination. This report thus reveals that BTB-A2s serve as negative regulators for balancing the intensity of ABA signaling during seed germination.

Although abscisic acid (ABA) is an important hormone that regulates seed dormancy, stomatal closure, plant development, as well as responses to environmental stimuli, the physiological mechanisms of ABA response to multiple stress in rice remain poorly understood. In the ABA biosynthetic pathway, 9-cis-epoxycarotenoid dioxygenase (NCED) is the key rate-limiting enzyme. Here, we report important functions of OsNCED3 in multi-abiotic stress tolerance in rice. The OsNCED3 is constitutively expressed in various tissues under normal condition, Its expression is highly induced by NaCl, PEG, and H2O2 stress, suggesting the roles for OsNCED3 in response to the multi-abiotic stress tolerance in rice. Compared with wild-type plants, nced3 mutants had earlier seed germination, longer post-germination seedling growth, increased sensitivity to water stress and H2O2 stress and increased stomata aperture under water stress and delayed leaf senescence. Further analysis found that nced3 mutants contained lower ABA content compared with wild-type plants, overexpression of OsNCED3 in transgenic plants could enhance water stress tolerance, promote leaf senescence and increase ABA content. We conclude that OsNCED3 mediates seed dormancy, plant growth, abiotic stress tolerance, and leaf senescence by regulating ABA biosynthesis in rice; and may provide a new strategy for improving the quality of crop.

Magnesium (Mg) is one of the essential nutrients for all living organisms. Plants acquire Mg from the environment and distribute within their bodies in the ionic form via Mg2+-permeable transporters. In Arabidopsis, the plasma membrane-localized magnesium transporter MGT6 mediates Mg2+ uptake under Mg-limited conditions, and therefore is important for the plant adaptation to low-Mg environment. In this study, we further assessed the physiological function of MGT6 using a knockout T-DNA insertional mutant allele. We found that MGT6 was required for normal plant growth during various developmental stages when the environmental Mg2+ was low. Interestingly, in addition to the hypersensitivity to Mg2+ limitation, mgt6 mutants displayed dramatic growth defects when external Mg2+ was in excess. Compared with wild-type plants, mgt6 mutants generally contained less Mg2+ under both low and high external Mg2+ conditions. Reciprocal grafting experiments further underpinned a role of MGT6 in a shoot-based mechanism for detoxifying excessive Mg2+ in the environment. Moreover, we found that mgt6 mgt7 double mutant showed more severe phenotypes compared with single mutants under both low- and high-Mg2+ stress conditions, suggesting that these two MGT-type transporters play an additive role in controlling plant Mg2+ homeostasis under a wide range of external Mg2+ concentrations.

Auxin controls a myriad of plant developmental processes and plant response to environmental conditions. Precise trafficking of auxin transporters is essential for auxin homeostasis in plants. Here, we report characterization of Arabidopsis CTL1, which controls seedling growth and apical hook development by regulating intracellular trafficking of PIN-type auxin transporters. The CTL1 gene encodes a choline transporter-like protein with an expression pattern highly correlated with auxin distribution and is enriched in shoot and root apical meristems, lateral root primordia, the vascular system, and the concave side of the apical hook. The choline transporter-like 1 (CTL1) protein is localized to the trans-Golgi network (TGN), prevacuolar compartment (PVC), and plasma membrane (PM). Disruption of CTL1 gene expression alters the trafficking of 2 auxin efflux transporters-Arabidopsis PM-located auxin efflux transporter PIN-formed 1 (PIN1) and Arabidopsis PM-located auxin efflux transporter PIN-formed 3 (PIN3)-to the PM, thereby affecting auxin distribution and plant growth and development. We further found that phospholipids, sphingolipids, and other membrane lipids were significantly altered in the ctl1 mutant, linking CTL1 function to lipid homeostasis. We propose that CTL1 regulates protein sorting from the TGN to the PM through its function in lipid homeostasis.

During double fertilization of angiosperms, the central cell of the female gametophyte fuses with a sperm cell to produce the endosperm, a storage tissue that nourishes the developing embryo within the seed. Although many genetic mutants defective in female gametophytic functions have been characterized, the molecular mechanisms controlling the specification and differentiation of the central cell are still not fully understood. Here, we report a mitochondrial ribosomal protein, RPS9M, is required for central cell maturation. RPS9M was highly expressed in the male and female gametophytes before and after double fertilization. The female gametophytes were defective in the rps9m mutant specifically concerning maturation of central cells. The morphological defects include unfused polar nuclei and smaller central vacuole in central cells. In addition, embryo initiation and early endosperm development were also severely affected in rps9m female gametophytes even after fertilized with wild type pollens. The RPS9M can interact with ANK6, an ankyrin-repeat protein in mitochondria previously reported to be required for fertilization. The expression pattern and mutant phenotype of RPS9M are similar to those of ANK6 as well, suggesting that RPS9M may work together with ANK6 in controlling female gametophyte development, possibly by regulating the expression of some mitochondrial proteins.

Rice (Oryza sativa; background Nipponbare) contains nine HKT (high-affinity K+ transport)-like genes encoding membrane proteins belonging to the superfamily of Ktr/TRK/HKT. OsHKTs have been proposed to include four selectivity filter-pore-forming domains homologous to the bacterial K+ channel KcsA, and are separated into OsHKT1s with Na+-selective activity and OsHKT2s with Na+-K+ symport activity. As a member of the OsHKT2 subfamily, OsHKT2;4 renders Mg2+ and Ca2+ permeability for yeast cells and Xenopus laevis oocytes, besides K+ and Na+. However, physiological functions related to Mg2+in planta have not yet been identified. Here we report that OsHKT2;4 from rice (O. sativa; background Nipponbare) functions as a low-affinity Mg2+ transporter to mediate Mg2+ homeostasis in plants under high-Mg2+ environments. Using the functional complementation assay in Mg2+-uptake deficient Salmonella typhimurium strains MM281 and electrophysiological analysis in X. laevis oocytes, we found that OsHKT2;4 could rescue the growth of MM281 in Mg2+-deficient conditions and induced the Mg2+ currents in oocytes at millimolar range of Mg2+. Additionally, overexpression of OsHKT2;4 to Arabidopsis mutant lines with a knockout of AtMGT6, a gene encoding the transporter protein necessary for Mg2+ adaptation in Arabidopsis, caused the Mg2+ toxicity to the leaves under the high-Mg2+ stress, but not under low-Mg2+ environments. Moreover, this Mg2+ toxicity symptom resulted from the excessive Mg2+ translocation from roots to shoots, and was relieved by the increase in supplemental Ca2+. Together, our results demonstrated that OsHKT2;4 is a low-affinity Mg2+ transporter responsible for Mg2+ transport to aerials in plants under high-Mg2+ conditions.

Abscisic acid (ABA) has been known participate in a wider range of adaptive responses to diverse environmental abiotic stresses such as drought, osmosis, and low temperatures. ABA signaling is initiated by its receptors PYR/PYL/RCARs, a type of soluble proteins with a conserved START domain which can bind ABA and trigger the downstream pathway. Previously, we discovered that poplar (Populus trichocarpa) genome encodes 14 PYR/PYL/RCAR orthologs (PtPYRLs), and two of them, PtPYRL1 and PtPYRL5 have been functionally characterized to positively regulate drought tolerance. However, the physiological function of these ABA receptors in poplar remains uncharacterized. Here, we generated transgenic poplar plants overexpressing PtPYRL1 and PtPYRL5 and found that they exhibited more vigorous growth and produced greater biomass when exposed to drought stress. The improved drought tolerance was positively correlated with the key physiological responses dictated by the ABA signaling pathway, including increase in stomatal closure and decrease in leaf water loss. Further analyses revealed that overexpression lines showed improved capacity in scavenging reactive oxygen species and enhanced the activation of antioxidant enzymes under drought stress. Moreover, overexpression of PtPYRL1 or PtPYRL5 significantly increased the poplar resistance to osmotic and cold stresses. In summary, our results suggest that constitutive expression of PtPYRL1 and PtPYRL5 significantly enhances the resistance to drought, osmotic and cold stresses by positively regulating ABA signaling in poplar.

Regarding the practical farming of Litopenaeus vannamei, the deterioration of water quality from intensive culture systems and environmental pollution is a common but troublesome problem in the cultivation of this species. The toxicities that result from deteriorating water quality, such as that from ammonia stress, have lethal effects on juvenile shrimp and can increase their susceptibility to pathogens. The toxicity of ammonia plays an important role in the frequently high mortality during the early stage on shrimp farms. However, little information is available regarding the genetic parameters of the ammonia tolerance of juveniles in the early stage, but this information is necessary to understand the potential for the genetic improvement of this trait. Considering the euryhalinity of L. vannamei and the fact that low salinity can increase the toxicity of ammonia stress, we estimated the heritability of ammonia tolerance in juveniles in 30‰ (normal) and 5‰ (low) salinity in this study using the survival time (ST) at individual level and the survival status at the half-lethal time (SS50) at the family level. In the normal and low salinity conditions and for the merged data, the heritability estimates of the ST (0.784±0.070, 0.575±0.068, and 0.517±0.058, respectively) and SS50 (0.402±0.061, 0.216±0.050, and 0.264±0.050, respectively) were all significantly greater than zero, which indicates that the ammonia-tolerance of shrimp can be greatly improved. So it might provide an alternative method to reduce mortality, help to enhance resistance to pathogens and reduce the occurrence of infectious diseases. The significant positive genetic correlation between ST and body length suggested that ammonia is more toxic to shrimp in the early stage. The medium-strength genetic correlations of the ST and SS50 between the two environments (0.394±0.097 and 0.377±0.098, respectively) indicate a strong genotype-by-environment (G×E) interaction for ammonia tolerance between the different salinity levels. Therefore, salinity-specific breeding programs for ammonia tolerance in shrimp should be purposefully implemented.

In this study, genetic parameters were obtained for growth and cold tolerance of 99 Fenneropenaeus chinensis juvenile families by means of indoor artificial cooling (starting from 14°C, 2°C/d). A linear mixed model was fitted to estimate variance components using the ASReml software package. Heritabilities estimated for body weight (BW) and body length (BL) of F. chinensis juveniles were 0.078 ± 0.124 and 0.131 ± 0.133, respectively. The estimates of heritability were low in magnitude for both traits. The differences between the heritabilities estimated for the two growth traits were not significant with each other, and the heritabilities were not significantly different from zero (P > 0.05). The phenotypic and genetic correlation coefficients between BW and BL were as high as 0.9408 ± 0.0040 and 0.9562 ± 0.0551, respectively, and both were significantly different from zero (P < 0.01). The heritabilities of temperature at death (TAD) and cooling degree hours (CDH) were 0.265 ± 0.091 and 0.077 ± 0.058, respectively. The heritability estimates for TAD was moderate and significantly different from zero (P < 0.01). The correlation analysis indicated that the phenotypic correlation coefficient between TAD and CDH was -0.5470 ± 0.0174, and the genetic correlation coefficient was -0.6707 ± 0.3635. In the analysis of growth traits and cold tolerance traits, the values of phenotypic correlation coefficient were floating between -0.1055 and 0.1098, both were significantly different from zero (P < 0.05), while the genetic correlation had a larger range (0.0526 ~ 0.9914), and all were not significantly different from zero (P > 0.05). In this study, there was a low correlation between growth and cold tolerance traits, indicating that growth traits and cold tolerance traits should be considered collectively in the breeding program of shrimp.

Although arsenite [As(III)] is non-essential and toxic for plants, it is effectively absorbed through various transporters into the roots. Here we identified a calcium-dependent protein kinase (CPK31) response for As(III) tolerance in Arabidopsis. We identified CPK31 as an interacting protein of a nodulin 26-like intrinsic protein (NIP1;1), an aquaporin involved in As(III) uptake. Similarly to the nip1;1 mutants, the loss-of-function mutants of CPK31 improved the tolerance against As(III) but not As(V), and accumulated less As(III) in roots than that of the wild-type plants. The promoter-β-glucuronidase and quantitative Real-Time PCR analysis revealed that CPK31 displayed overlapping expression profiles with NIP1;1 in the roots, suggesting that they might function together in roots. Indeed, the cpk31 nip1;1 double mutants exhibited stronger As(III) tolerance than cpk31 mutants, but similar to nip1;1 mutants, supporting the idea that CPK31 might serve as an upstream regulator of NIP1;1. Furthermore, transient CPK31 overexpression induced by dexamethasone caused the decrease in As(III) tolerance of transgenic Arabidopsis lines. These findings reveal that CPK31 is a key factor in As(III) response in plants.

In Arabidopsis, the salt overly sensitive (SOS) pathway, consisting of calcineurin B-like protein 4 (CBL4/SOS3), CBL-interacting protein kinase 24 (CIPK24/SOS2) and SOS1, has been well defined as a crucial mechanism to control cellular ion homoeostasis by extruding Na+ to the extracellular space, thus conferring salt tolerance in plants. CBL10 also plays a critical role in salt tolerance possibly by the activation of Na+ compartmentation into the vacuole. However, the functional relationship of the SOS and CBL10-regulated processes remains unclear. Here, we analyzed the genetic interaction between CBL4 and CBL10 and found that the cbl4 cbl10 double mutant was dramatically more sensitive to salt as compared to the cbl4 and cbl10 single mutants, suggesting that CBL4 and CBL10 each directs a different salt-tolerance pathway. Furthermore, the cbl4 cbl10 and cipk24 cbl10 double mutants were more sensitive than the cipk24 single mutant, suggesting that CBL10 directs a process involving CIPK24 and other partners different from the SOS pathway. Although the cbl4 cbl10, cipk24 cbl10, and sos1 cbl10 double mutants showed comparable salt-sensitive phenotype to sos1 at the whole plant level, they all accumulated much lower Na+ as compared to sos1 under high salt conditions, suggesting that CBL10 regulates additional unknown transport processes that play distinct roles from the SOS1 in Na+ homeostasis.

Magnesium (Mg2+) is an essential nutrient in all organisms. However, high levels of Mg2+ in the environment are toxic to plants. In this study, we identified the vacuolar-type H+-pyrophosphatase, AVP1, as a critical enzyme for optimal plant growth under high-Mg conditions. The Arabidopsis avp1 mutants displayed severe growth retardation, as compared to the wild-type plants upon excessive Mg2+. Unexpectedly, the avp1 mutant plants retained similar Mg content to wild-type plants under either normal or high Mg conditions, suggesting that AVP1 may not directly contribute to Mg2+ homeostasis in plant cells. Further analyses confirmed that the avp1 mutant plants contained a higher pyrophosphate (PPi) content than wild type, coupled with impaired vacuolar H+-pyrophosphatase activity. Interestingly, expression of the Saccharomyces cerevisiae cytosolic inorganic pyrophosphatase1 gene IPP1, which facilitates PPi hydrolysis but not proton translocation into vacuole, rescued the growth defects of avp1 mutants under high-Mg conditions. These results provide evidence that high-Mg sensitivity in avp1 mutants possibly resulted from elevated level of cytosolic PPi. Moreover, genetic analysis indicated that mutation of AVP1 was additive to the defects in mgt6 and cbl2 cbl3 mutants that are previously known to be impaired in Mg2+ homeostasis. Taken together, our results suggest AVP1 is required for cellular PPi homeostasis that in turn contributes to high-Mg tolerance in plant cells.

Drought stress is an important environmental factor limiting productivity of plants, especially fast growing species with high water consumption like poplar. Abscisic acid (ABA) is a phytohormone that positively regulates seed dormancy and drought resistance. The PYR1 (Pyrabactin Resistance 1)/ PYRL (PYR-Like)/ RCAR (Regulatory Component of ABA Receptor) (PYR/PYL/RCAR) ABA receptor family has been identified and widely characterized in Arabidopsis thaliana. However, their functions in poplars remain unknown. Here, we report that 2 of 14 PYR/PYL/RCAR orthologues in poplar (Populus trichocarpa) (PtPYRLs) function as a positive regulator of the ABA signal transduction pathway. The Arabidopsis transient expression and yeast two-hybrid assays showed the interaction among PtPYRL1 and PtPYRL5, a clade A protein phosphatase 2C, and a SnRK2, suggesting that a core signalling complex for ABA signaling pathway exists in poplars. Phenotypic analysis of PtPYRL1 and PtPYRL5 transgenic Arabidopsis showed that these two genes positively regulated the ABA responses during the seed germination. More importantly, the overexpression of PtPYRL1 and PtPYRL5 substantially improved ABA sensitivity and drought stress tolerance in transgenic plants. In summary, we comprehensively uncovered the properties of PtPYRL1 and PtPYRL5, which might be good target genes to genetically engineer drought-Resistant plants.

In the practical farming of Litopenaeus vannamei, the intensive culture system and environmental pollution usually results in a high concentration of ammonia, which usually brings large detrimental effects to shrimp, such as increasing the susceptibility to pathogens, reducing growth, decreasing osmoregulatory capacity, increasing the molting frequency, and even causing high mortality. However, little information is available on the molecular mechanisms of the detrimental effects of ammonia stress in shrimp. In this study, we performed comparative transcriptome analysis between ammonia-challenged and control groups from the same family of L. vannamei to identify the key genes and pathways response to ammonia stress. The comparative transcriptome analysis identified 136 significantly differentially expressed genes that have high homologies with the known proteins in aquatic species, among which 94 genes are reported potentially related to immune function, and the rest of the genes are involved in apoptosis, growth, molting, and osmoregulation. Fourteen GO terms and 6 KEGG pathways were identified to be significantly changed by ammonia stress. In these GO terms, 13 genes have been studied in aquatic species, and 11 of them were reported potentially involved in immune defense and two genes were related to molting. In the significantly changed KEGG pathways, all the 7 significantly changed genes have been reported in shrimp, and four of them were potentially involved in immune defense and the other three were related to molting, defending toxicity, and osmoregulation, respectively. In addition, majority of the significantly changed genes involved in nitrogen metabolisms that play an important role in reducing ammonia toxicity failed to perform the protection function. The present results have supplied molecular level support for the previous founding of the detrimental effects of ammonia stress in shrimp, which is a prerequisite for better understanding the molecular mechanism of the immunosuppression from ammonia stress.