Search

Summary: MicroRNAs (miRNAs) play crucial roles in regulating plant development and stress responses. However, the functions and mechanism of intronic miRNAs in plants are poorly understood.This study reports a stress‐responsive RNA splicing mechanism for intronic miR400 production, whereby miR400 modulates reactive oxygen species (ROS) accumulation and improves plant tolerance by downregulating its target expression.To monitor the intron splicing events, we used an intronic miR400 splicing‐dependent luciferase transgenic line. Luciferase activity was observed to decrease after high cadmium concentration treatment due to the retention of the miR400‐containing intron, which inhibited the production of mature miR400. Furthermore, we demonstrated that under Cd treatments, Pentatricopeptide Repeat Protein 1 (PPR1), the target of miR400, acts as a positive regulator by inducing ROS accumulation. Ppr1 mutation affected the Complex III activity in the electron transport chain and RNA editing of the mitochondrial gene ccmB.This study illustrates intron splicing as a key step in intronic miR400 production and highlights the function of intronic miRNAs as a 'signal transducer' in enhancing plant stress tolerance. [ABSTRACT FROM AUTHOR]

Increasing evidence points to the tight relationship between alternative splicing (AS) and the salt stress response in plants. However, the mechanisms linking these two phenomena remain unclear. In this study, we have found that Salt-Responsive Alternatively Spliced gene 1 (SRAS1), encoding a RING-Type E3 ligase, generates two splicing variants: SRAS1.1 and SRAS1.2, which exhibit opposing responses to salt stress. The salt stress-responsive AS event resulted in greater accumulation of SRAS1.1 and a lower level of SRAS1.2. Comprehensive phenotype analysis showed that overexpression of SRAS1.1 made the plants more tolerant to salt stress, whereas overexpression of SRAS1.2 made them more sensitive. In addition, we successfully identified the COP9 signalosome 5A (CSN5A) as the target of SRAS1. CSN5A is an essential player in the regulation of plant development and stress. The full-length SRAS1.1 promoted degradation of CSN5A by the 26S proteasome. By contrast, SRAS1.2 protected CSN5A by competing with SRAS1.1 on the same binding site. Thus, the salt stress-triggered AS controls the ratio of SRAS1.1/SRAS1.2 and switches on and off the degradation of CSN5A to balance the plant development and salt tolerance. Together, these results provide insights that salt-responsive AS acts as post-transcriptional regulation in mediating the function of E3 ligase. Author summary: High salinity severely affects plant growth and development, impairing crop production worldwide. E3 ligase is a stress-responsive regulator through ubiquitin-proteasome system for selective protein degradation. The E3s are regulated by transcriptional regulation and post-translational modifications. Here, we have discovered that stress-responsive AS acts as a post-transcriptional regulation modulating the function of E3 ligases. Intriguingly, the truncated proteins generated by salt-responsive AS play opposite roles compared with the full-length E3 ligase. The truncated isoform losing key domain could not degrade the target protein, instead, it interacts and competes with the E3 ligase through binding the same domain of the targets. This finding contributes significantly to a deeper mechanistic understanding of how AS regulates the function of E3 ligase in response to salt stress. [ABSTRACT FROM AUTHOR]

Summary: MicroRNAs (miRNAs) have emerged as a class of regulators of gene expression through posttranscriptional degradation or translational repression in living cells. Increasing evidence points to the important relationship between miRNAs and environmental stress responses, but the regulatory mechanisms in plants are poorly understood. Here, we found that Arabidopsis thaliana intronic miR400 was cotranscribed with its host gene (At1g32583) and downregulated by heat treatment. Intriguingly, an alternative splicing (AS) event that occurred in the intron (306 bp) where MIR400 was located was specifically induced by heat stress. A 100 bp fragment was excised, and the remaining 206 bp intron containing MIR400 transcripts was retained in the host gene. The stress-induced AS event thus resulted in greater accumulation of miR400 primary transcripts and a low level of mature miR400. Together, these results provide the direct evidence that AS acts as a regulatory mechanism linking miRNAs and environmental stress in plants. [Copyright &y& Elsevier]

Numerous studies have demonstrated the function of salinity or jasmonic acid (JA) in plant growth and senescence. This study evaluated how the combination of salinity and methyl jasmonate (MeJA) (SaM) worked as a novel stress and then regulated plant growth in Arabidopsis . Firstly, we found that compared with MeJA or NaCl treatment alone, SaM would significantly intensified plant growth inhibition and senescence in wild-type (WT) seedlings, and these phenotypes could be partially compromised after SaM stress in JA-insensitive mutants. Meanwhile, genes involved in JA signaling and Senescence Associated Gene 13 ( SAG13 ) were dramatically increased by SaM stress than that by MeJA or NaCl alone in WT. Moreover, a group of secondary metabolite − indolic glucosinolates (IGs) showed obvious over-accumulation after SaM treatment than that after each single one in WT, and the seedlings treated with IGs’ metabolites performed similar inhibited growth and chlorotic leaves phenotypes compared with those caused by SaM stress. All these indicated the toxicity of IGs and their metabolites would prevent the growth progress of plants. Therefore, we concluded that SaM worked as a novel stress and intensified plant growth inhibition and senescence, which was dependent on JA-dependent and −independent signaling pathways. [ABSTRACT FROM AUTHOR]

Mitogen-activated protein kinase ( MAPK) cascades are involved in plant development, stress responses and hormonal signal transduction. MAPK kinases ( MAPKKs), as the key nodes in these cascades, link MAPKs and MAPKK kinases ( MAPKKKs). In this study, Gh MKK4, a novel group C MAPKK gene from cotton ( Gossypium hirsutum), was isolated and identified. Its expression can be induced by various stresses and signalling molecules. The overexpression of GhMKK4 in Nicotiana benthamiana enhanced its susceptibility to bacterial and fungal pathogens, but had no significant effects on salt or drought tolerance. Notably, the overexpressing plants showed increased sensitivity to abscisic acid ( ABA) and gibberellin A3 ( GA3), and ABA and gibberellin ( GA) signalling were affected on infection with Ralstonia solanacearum bacteria. Furthermore, the overexpressing plants showed more reactive oxygen species ( ROS) accumulation and stronger inhibition of catalase ( CAT), a ROS-scavenging enzyme, than control plants after salicylic acid ( SA) treatment. Interestingly, two genes encoding ornithine decarboxylase ( ODC) and S-adenosylmethionine decarboxylase ( SAMDC), the key enzymes in polyamine synthesis, exhibited reduced R. solanacearum-induced expression in overexpressing plants. These findings broaden our knowledge about the functions of MAPKKs in diverse signalling pathways and the negative regulation of disease resistance in the cotton crop. [ABSTRACT FROM AUTHOR]

The nuclear factor Y (NF-Y), which is a ubiquitous transcription factor found in eukaryotes, is composed of three distinct subunits, namely, NF-YA, NF-YB, and NF-YC. Here, we firstly characterized the detailed function of the Arabidopsis NFYA1 factor. It is found that the 35S::AtNFYA1-overexpressed lines were hypersensitive to salt stress and Abscisic acid (ABA) during the early-postgermination growth stages. The transgenic lines exhibited a severe postgermination growth arrest compared with the wild-type (WT) under salt stress and ABA treatment. Interestingly, sodium tungstate, which is an ABA synthesis inhibitor, restored the salt-sensitive phenotype of the 35S::AtNFYA1 lines. Results of the qRT-PCR analysis showed that the mRNA levels of ABI3 and ABI5, as well as their downstream genes AtEM1 and AtEM6, were more greatly upregulated under salt stress during seed germination in the transgenic lines compared with those in WT. On the other hand, the NFYA1-RNAi lines were found to be insensitive to salt stress and exhibited decreased levels of ABI3, ABI5, EM1, and EM6 transcripts. Our results provide clear evidence supporting a role of AtNFYA1 in regulating postgermination growth arrest under salt stress. [ABSTRACT FROM AUTHOR]

Abstract: To characterize the microRNAs that contribute to the development of brace root, Solexa high-throughput sequencing of three libraries derived from tissues of node (N), nodes with just-emerged brace roots (NR), and nodes with just-emerged brace roots after IAA treatment (NRI) was performed. Total 650,793, 957,303 and 1,082,948 genome-matched unique reads were obtained in N, NR and NRI libraries, respectively. Further analysis confirmed the authenticity of 137 known miRNAs and the discovery of 159 novel miRNAs in maize. 14 conserved and 16 novel miRNAs differentially expressed in brace root, as well as 15 target genes, were identified and validated by qRT-PCR during maize brace root development. Moreover, we identified 9 miRNA precursor-matched novel sRNAs that may form miRNA clusters, as well as 24nt siRNAs in the three libraries. In addition, we suggest that auxin represent a regulator in brace root development and can be regulated at the posttranscriptional level by miRNAs. [Copyright &y& Elsevier]

Mitogen-activated protein kinase (MAPK) cascades are involved in various processes from plant growth and development to biotic and abiotic stress responses. MAPK kinases (MAPKKs), which link MAPKs and MAPKK kinases (MAPKKKs), play crucial roles in MAPK cascades to mediate a variety of stress responses in plants. However, few MAPKKs have been functionally characterized in cotton (Gossypium hirsutum). In this study, a novel gene, GhMKK5, from cotton belonging to the group C MAPKKs was isolated and characterized. The expression of GhMKK5 can be induced by pathogen infection, abiotic stresses, and multiple defence-related signal molecules. The overexpression of GhMKK5 in Nicotiana benthamiana enhanced the plants’ resistance to the bacterial pathogen Ralstonia solanacearum by elevating the expression of pathogen resistance (PR) genes, including PR1a, PR2, PR4, PR5, and NPR1, but increased the plants’ sensitivity to the oomycete pathogen Phytophthora parasitica var. nicotianae Tucker. Importantly, GhMKK5-overexpressing plants displayed markedly elevated expression of reactive oxygen species-related and cell death marker genes, such as NtRbohA and NtCDM, and resulted in hypersensitive response (HR)-like cell death characterized by the accumulation of H2O2. Furthermore, it was demonstrated that GhMKK5 overexpression in plants reduced their tolerance to salt and drought stresses, as determined by statistical analysis of seed germination, root length, leaf water loss, and survival rate. Drought obviously accelerated the cell death phenomenon in GhMKK5-overexpressing plants. These results suggest that GhMKK5 may play an important role in pathogen infection and the regulation of the salt and drought stress responses in plants. [ABSTRACT FROM PUBLISHER]

Abstract: Fructose 1,6-biphosphate aldolase (FBA) is a key enzyme in plants, which is involved not only in glycolysis and gluconeogenesis in the cytoplasm, but also in the Calvin cycle in plastids. Research on FBAs in various organisms has been reported, but there is none on FBAs in Arabidopsis at the molecular level. In the current study, eight FBA family genes (AtFBA1–8) were identified and analyzed in Arabidopsis thaliana. These genes have a highly conserved aldolase-type TIM barrel domain and a C-terminal peptide, but variable N-terminal peptides. Based on the phylogenetic analysis of FBA protein sequences from Arabidopsis and other plant species, AtFBA family was classified into two subfamilies, including three members (AtFBA1–3) with high similarities to FBAs occurring at plastid, and five (AtFBA4–8) with high similarities to FBAs localized in the cytoplasm. By confocal microscopy analysis with GFP fusion protein, AtFBA3 and AtFBA4 as well as AtFBA6 were observed to be localized in the plastid and cytoplasm, respectively. At least two duplicated gene pairs of AtFBA1 and AtFBA2, as well as AtFBA4 and AtFBA8 were found. Transcript level analysis of AtFBA genes in various tissues revealed the unique and overlapping expression patterns of plastid and cytosol AtFBA genes, suggesting that these genes may function at different stages of plant growth and development. Interestingly, AtFBA1, AtFBA2, AtFBA5 and AtFBA7 showed undetectable expression in roots. The expression patterns of AtFBA genes under different stress conditions suggested that all the members showed different expression patterns in response to stresses, including ABA, NaCl, Cd, abnormal temperature and drought, and, except for AtFBA3, most of the AtFBA genes were significantly responsive to drought stress in roots. Moreover, AtFBA1, AtFBA2, AtFBA5, AtFBA7 and AtFBA8 were induced by at least one of three sugars (sucrose, glucose and fructose) after 24h of treatment. Further functional analyses indicated important clues of AtFBA2, AtFBA6 and AtFBA8 in plant growth, stress responses and development, respectively. Thus these results provide additional knowledge on AtFBA families and their roles. [Copyright &y& Elsevier]

The molecular mechanism for sensing and transducing the stress signals initiated by K deprivation in plants remains unknown. Here, we found that the expression of AtHELPS, an Arabidopsis DExD/H box RNA helicase gene, was induced by low-K, zeatin and cold treatments, and downregulated by high-K stress. To further investigate the expression pattern of AtHELPS, pAtHELPS::GUS transgenic plants were generated. Histochemical staining indicated that AtHELPS is mainly expressed in the young seedlings and vascular tissues of leaves and roots. Using both helps mutants and overexpression lines, we observed that, in the low-K condition, AtHELPS affected Arabidopsis seed germination and plant weight. Interestingly, the mRNA levels of AKT1, CBL1/9 and CIPK23 in the helps mutants were much higher than in the overexpression lines under low-K stress. Moreover, under low-K stress, the helps mutants displayed increased K influx, whereas the overexpression line of AtHELPS had a lower flux rate in the roots by the noninvasive micro-test technique. Taken together, these results provide information for the functional analysis of plant DEVH box RNA helicases, and suggest that AtHELPS, as an important negative regulator, plays a role in K deprivation stress. The Arabidopsis DExD/H box RNA helicase gene, AtHELPS, was mainly expressed in young seedlings and vascular tissues of leaves and roots. It was also induced by low potassium stress and down-regulated by high potassium stress. In the low potassium condition AtHELPS affected Arabidopsis seed germination and plant weight by affecting transcription of AKT1, CBL1/9 and CIPK23 and K influx rate [ABSTRACT FROM AUTHOR]

Mitogen-activated protein kinase (MAPK) cascades play important roles in mediating pathogen responses and reactive oxygen species signaling. In plants, MAPKs are classified into four major groups (A-D). Previous studies have mainly focused on groups A and B, but little is known about group C. In this study, we functionally characterized a stress-responsive group C MAPK gene ( GhMPK2) from cotton. Northern blot analysis indicated that GhMPK2 was induced not only by signaling molecules, such as ethylene and methyl jasmonate, but also by methyl viologen-mediated oxidative stress. Transgenic tobacco ( Nicotiana tabacum) plants that overexpress GhMPK2 displayed enhanced resistance to fungal and viral pathogens, and the expression of the pathogenesis-related (PR) genes, including PR1, PR2, PR4, and PR5, was significantly increased. Interestingly, the transcription of 1-aminocyclopropane-1-carboxylic acid synthase ( ACS) and 1-aminocyclopropane-1-carboxylic acid oxidase ( ACO) was significantly upregulated in transgenic plants, suggesting that GhMPK2 positively regulates ethylene synthesis. Moreover, overexpression of GhMPK2 elevated the expression of several antioxidant enzymes, conferring on transgenic plants enhanced reactive oxygen species scavenging capability and oxidative stress tolerance. These results increased our understanding of the role of the group C GhMPK2 gene in multiple defense-signaling pathways, including those that are involved in responses to pathogen infection and oxidative stress. In this study, functional characterization of a cotton group C mitogen-activated protein kinase (MAPK) gene ( GhMPK2) was carried out. The expression of GhMPK2 was induced by signaling molecules and oxidative stress. Overexpression of GhMPK2 significantly elevated the expression of the pathogenesis-related ( PR), signaling-related and antioxidative genes, and thereby enhanced the transgenic plants disease resistance and oxidative stress tolerance. [ABSTRACT FROM AUTHOR]

Currently, the molecular regulation mechanisms involved in the early development of maize brace root are poorly known. To gain insight into the transcriptome dynamics that are associated with its development, genome-wide gene expression profiling was conducted by Solexa sequencing (Illumina Inc., San Diego, CA, USA). More than five million tags were generated from the stem node tissues without and with just-emerged brace roots, including 149 524 and 178 131 clean tags in the two libraries, respectively. Of these, 82 864 (55.4%) and 91 678 (51.5%) tags were matched to the reference genes. The most differentially expressed tags with a log ratio > 2 or < −2 ( P < 0.001) were analyzed further, representing 143 up-regulated and 152 down-regulated genes, except for unknown transcripts, which were classified into 11 functional categories. The most enriched categories were those of metabolism, signal transduction and cellular transport. Many genes or biological pathways were found to be commonly shared between brace root and lateral or adventitious root development, such as genes participating in cell wall degradation and synthesis, auxin transport and signaling, ethylene signaling, etc. Next, the expression patterns of 20 genes were assessed by quantitative real-time PCR, and the results obtained showed general agreement with the Solexa analysis. Furthermore, a comparison of the brace root transcriptome with that of maize primary root revealed substantial differences in the categories and abundances of expressed transcripts. In conclusion, we first reveal the complex changes in the transcriptome during the early development of maize brace root and provide a comprehensive set of data that are essential for understanding its molecular regulation. [ABSTRACT FROM AUTHOR]

Enzymatic and non-enzymatic antioxidants play important roles in the tolerance of abiotic stress. To increase the resistance of seeds to oxidative stress, At2S3 promoter from Arabidopsis was used to achieve overexpression of the antioxidants in a seed-specific manner. This promoter was shown to be capable of driving the target gene to have a high level of expression in seed-related organs, including siliques, mature seeds, and early seedlings, thus making its molecular farming applications in plants possible. Subsequently, genes encoding Mn-superoxide dismutase ( MSD1), catalase ( CAT1), and homogentisate phytyltransferase ( HPT1, responsible for the first committed reaction in the tocopherol biosynthesis pathway) were overexpressed in Arabidopsis under the control of the At2S3 promoter. Double overexpressers co-expressing two enzymes and triple overexpressers were produced by cross pollination. Mn-SOD and total CAT activities, as well as γ-tocopherol content, significantly increased in the corresponding overproduction lines. Moreover, single MSD1-transgene, double, and triple overexpressers displayed remarkably enhanced oxidative stress tolerance compared to wild type during seed germination and early seedling growth. Interestingly, an increase in the total CAT activity was also observed in the single MSD1-transgenic lines as a result of MSD1 overexpression. Together, the combined increase in Mn-SOD and CAT activities in seeds plays an essential role in the improvement of antioxidant capacity at early developmental stage in Arabidopsis. [ABSTRACT FROM AUTHOR]

Abstract: Being poikilothermic and sessile organisms, plants have to respond quickly to changing environmental cues, and a higher order of gene regulation is required. The significance of nucleocytoplasmic transport via importinα and importinβ (α/β) has been exhibited in a wide spectrum of biological processes. However, most of these receptors have not been characterized as to which cellular or development processes are required and how their expression is regulated by environmental stimuli. Here we pursued a phylogenetic analysis and investigated the expression patterns of all 8 IMPαs and 18 IMPβs in Arabidopsis. The IMPα isoforms could be tracked back to a common ancestor, while the IMPβs derived from different ones. The majority of transport receptor genes were constitutively expressed. Intriguingly, AtIMPα5, 7, 8 and AtIMPβ5 were specifically expressed in different tissues. AtIMPβ3 was the sole receptor that was obviously modulated by exogenous phytohormones, whereas three IMPαs and five IMPβs exhibited responses to environmental stimuli. Furthermore, our RT-PCR data provided direct evidence that AtIMPα5, 8 and AtIMPβ5 are not pseudogenes and we also corrected the open reading frame annotation of AtIMPα8. These genome-wide profiling results not only widen our understanding of these transport receptors, but also provide strong evidence supporting the role of transport receptors in multiple signaling pathways and give us an insight into the further analysis of nucleocytoplasmic trafficking in Arabidopsis. [Copyright &y& Elsevier]