Search

Pre-harvest sprouting (PHS) adversely affects wheat quality and yield, and grain color (GC) is associated with PHS resistance. However, the genetic relationship between GC and PHS resistance remains unclear. In this study, 168 wheat varieties (lines) with significant differences in GC and PHS resistance were genotyped using an Illumina 90K iSelect SNP array. Genome-wide association study (GWAS) based on a mixed linear model showed that 67 marker-trait associations (MTAs) assigned to 29 loci, including 17 potentially novel loci, were significantly associated with GC, which explained 1.1–17.0% of the phenotypic variation. In addition, 100 MTAs belonging to 54 loci, including 31 novel loci, were significantly associated with PHS resistance, which accounted for 1.1–14.7% of the phenotypic variation. Subsequently, two cleaved amplified polymorphic sequences (CAPS) markers, 2B-448 on chromosome 2B and 5B-301 on chromosome 5B, were developed from the representative SNPs of the major common loci Qgc.ahau-2B.3/Qphs.ahau-2B.4 controlling GC/PHS resistance and PHS resistance locus Qphs.ahau-5B.4, respectively. Further validation in 171 Chinese mini-core collections confirmed significant correlations of the two CAPS markers with GC and PHS resistance phenotypes under different environments (P

IntroductionSeed dormancy (SD) significantly decreases under high temperature (HT) environment during seed maturation, resulting in pre-harvest sprouting (PHS) damage under prolonged rainfall and wet weather during wheat harvest. However, the molecular mechanism underlying HT-mediated SD remains elusiveSeed dormancy (SD) significantly decreases under high temperature (HT) environment during seed maturation, resulting in pre-harvest sprouting (PHS) damage under prolonged rainfall and wet weather during wheat harvest. However, the molecular mechanism underlying HT-mediated SD remains elusive.MethodsHere, the wheat landrace ‘Waitoubai’ with strong SD and PHS resistance was treated with HT from 21 to 35 days post anthesis (DPA). Then, the seeds under HT and normal temperature (NT) environments were collected at 21 DPA, 28 DPA, and 35 DPA and subjected to whole-transcriptome sequencing.ResultsThe phenotypic data showed that the seed germination percentage significantly increased, whereas SD decreased after HT treatment compared with NT, consistent with the results of previous studies. In total, 5128 mRNAs, 136 microRNAs (miRNAs), 273 long non-coding RNAs (lncRNAs), and 21 circularRNAs were found to be responsive to HT, and some of them were further verified through qRT-PCR. In particular, the known gibberellin (GA) biosynthesis gene TaGA20ox1 (TraesCS3D02G393900) was proved to be involved in HT-mediated dormancy by using the EMS-mutagenized wheat cultivar Jimai 22. Similarly, a novel gene TaCDPK21 (TraesCS7A02G267000) involved in the calcium signaling pathway was validated to be associated with HT-mediated dormancy by using the EMS mutant. Moreover, TaCDPK21 overexpression in Arabidopsis and functional complementarity tests supported the negative role of TaCDPK21 in SD. We also constructed a co-expression regulatory network based on differentially expressed mRNAs, miRNAs, and lncRNAs and found that a novel miR27319 was located at a key node of this regulatory network. Subsequently, using Arabidopsis and rice lines overexpressing miR27319 precursor or lacking miR27319 expression, we validated the positive role of miR27319 in SD and further preliminarily dissected the molecular mechanism of miR27319 underlying SD regulation through phytohormone abscisic acid and GA biosynthesis, catabolism, and signaling pathways.DiscussionThese findings not only broaden our understanding of the complex regulatory network of HT-mediated dormancy but also provide new gene resources for improving wheat PHS resistance to minimize PHS damage by using the molecular pyramiding approach.

Genetic improvement of grain yield is always an important objective in wheat breeding. Here, a genome-wide association study was conducted to parse the complex genetic composition of yield-related traits of 105 elite wheat varieties (lines) using the Wheat 90K Illumina iSelect SNP array. Nine yield-related traits, including maximum number of shoots per square meter (MSN), effective number of spikes per square meter (ESN), percentage of effective spike (PES), number of kernels per spike (KPS), thousand-kernel weight (TKW), the ratio of kernel length/kernel width (RLW), leaf-area index (LAI), plant height (PH), and grain yield (GY), were evaluated across four environments. Twenty four highly significant marker-trait associations (MTAs) (P < 0.001) were identified for nine yield-related traits on chromosomes 1A, 1D, 2A (2), 3B, 4A (2), 4B, 5A (4), 5B (4), 5D, 6B (2), 7A (2), and 7B (3), explaining 10.86-20.27% of the phenotypic variations. Of these, four major loci were identified in more than three environments, including one locus for RLW (6B), one locus for TKW (7A), and two loci for PH (7B). A cleaved amplified polymorphic sequence (CAPS) marker Td99211 for TKW on chromosome 5A was developed and validated in both a natural population composed of 372 wheat varieties (lines) and a RIL population derived from the cross of Yangxiaomai × Zhongyou 9507. The CAPS marker developed can be directly used for marker-assisted selection in wheat breeding, and the major MTAs identified can provide useful information for fine-mapping of the target genes in future studies.

As the main pigment in photosynthesis, chlorophyll significantly affects grain filling and grain weight of crop. Cytokinin (CTK) can effectively increase chlorophyll content and chloroplast stability, but it is irreversibly inactivated by cytokinin oxidase (CKX). In this study, therefore, twenty-four pairs of primers were designed to identify variations of wheat CKX (Tackx) genes associated with flag leaf chlorophyll content after anthesis, as well as grain weight in 169 recombinant inbred lines (RIL) derived from Triticum aestivum Jing 411 × Hongmangchun 21. Results indicated variation of Tackx4, identified by primer pair T19-20, was proven to significantly associate with chlorophyll content and grain weight in the RIL population. Here, two Tackx4 patterns were identified: one with two co-segregated fragments (Tackx4-1/Tackx4-2) containing 618 bp and 620 bp in size (as in Jing 411), and another with no PCR product. The two genotypes were designated as genotype-A and genotype-B, respectively. Grain weight and leaf chlorophyll content at 5~15 days after anthesis (DAA) were significantly higher in genotype-A lines than those in genotype-B lines. Mapping analysis indicated Tackx4 was closely linked to Xwmc169 on chromosome 3AL, as well as co-segregated with a major quantitative trait locus (QTL) for both grain weight and chlorophyll content of flag leaf at 5~15 DAA. This QTL explained 8.9~22.3% phenotypic variations of the two traits across four cropping seasons. Among 102 wheat varieties, a third genotype of Tackx4 was found and designated as genotype-C, also having two co-segregated fragments, Tackx4-2 and Tackx4-3 (615bp). The sequences of three fragments, Tackx4-1, Tackx4-2, and Tackx4-3, showed high identity (>98%). Therefore, these fragments could be considered as different copies at Tackx4 locus on chromosome 3AL. The effect of copy number variation (CNV) of Tackx4 was further validated. In general, genotype-A contains both significantly higher grain weight and flag leaf chlorophyll content at 5~15 DAA than those in genotype-B and genotype-C, among 102 varieties under various environments.

Cytokinin oxidase (CKX) plays a crucial role in plant growth and development by reversibly inactivating cytokinin (CTK). Twenty-four primer pairs, designed from ESTs of the TaCKX genes family of common wheat, were used to identify their allelic variations associated with grain size, weight, and filling rate in 169 recombinant inbred lines (RIL) derived from Jing 411 × Hongmangchun 21. TaCKX6a02, a member of TaCKX gene family, amplified by primer pair T31-32, showed a close association with grain traits in this RIL population. Statistical analysis indicated that allelic variation of TaCKX6a02 had significant correlation with grain size, weight, and filling rate (GFR; P < 0.001) under varied environments. The TaCKX6a02-D1a allele from Jing411 significantly increased grain size, weight and grain filling rate, compared with TaCKX6a02-D1b from Hongmangchun 21. TaCKX6a02 was located on chromosome 3DS in the interval of Xbarc1119 and Xbarc1162, with a genetic distance of 1.4 cM. The location was further confirmed using Chinese Spring nulli-tetrasomic lines. A major QTL (quantitative trait locus) tightly linked to TaCKX6a02 was detected in the RIL population, explaining 17.1~38.2% of phenotype variations for grain size, weight, GFRmax and GFRmean in different environments. In addition, significant effects of variations of TaCKX6a02 on grain weight and GFR were further validated by association analysis among 102 wheat varieties in two cropping seasons. 12.8~35.1% of phenotypic variations were estimated for these genotypes. A novel 29-bp InDel behind the stop codon was detected by DNA sequence analysis between the two alleles of TaCKX6a02-D1. The gene-specific marker, TKX3D, was designed according to the novel variation, and can be used in marker-assisted selection (MAS) for grain size, weight, and GFR in common wheat.

Posttranslational modification of tubulin increases the dynamic complexity and functional diversity of microtubules. Acetylation of α-tubulin at Lys-40 is a highly conserved posttranslational modification that has been shown to improve the flexibility and resilience of microtubules. Here we studied the in vivo functions of α-tubulin acetylation by knocking-out Atat, the Drosophila α-tubulin acetyltransferase, and by mutating Lys-40 to Arg in α1-tubulin. We found a reduction in the dendritic arborization of larval class I dendritic arborization (da) neurons in both mutants. The dendritic developmental defects in atat mutants could be reversed by enhancing the stability of microtubules either through knocking down the microtubule severing protein Katanin 60 or through overexpressing tubulin-specific chaperone E, suggesting that α-tubulin deacetylation impairsed dendritic morphology by decreasing the stability of microtubules. Using time-lapse recordings, we found that atat and α1-tubulinK40R mutations dramatically increased the number of dendritic protrusions that were likely to be immature dendritic precursors. Finally, we showed that both Atat and α-tubulin acetylation were required in class I da neurons to control larval locomotion. These findings add novel insight into the current knowledge of the role of α-tubulin acetylation in regulating neuronal development and functions.

Thousand-grain weight (TGW) of wheat (Triticum aestivum L.) contributes significantly to grain yield. In the present study, a candidate gene associated with TGW was identified through specifc-locus amplified fragment sequencing (SLAF-seq) of DNA bulks of recombinant inbred lines (RIL) derived from the cross between Jing 411 and Hongmangchun 21. The gene was located on chromosome 7A, designated as TaTGW-7A with a complete genome sequence and an open reading frame (ORF). A single nucleotide polymorphism (SNP) was present in the first exon between two alleles at TaTGW-7A locus, resulting in a Val to Ala substitution, corresponding to a change from higher to lower TGW. Cleaved amplified polymorphic sequence (CAPS) (TGW7A) and InDel (TG9) markers were developed to discriminate the two alleles TaTGW-7Aa and TaTGW-7Ab for higher and lower TGW, respectively. A major QTL co-segregating with TaTGW-7A explained 21.7-27.1% of phenotypic variance for TGW in the RIL population across five environments. The association of TaTGW-7A with TGW was further validated in a natural population and Chinese mini-core collections. Quantitative real-time PCR revealed higher transcript levels of TaTGW-7Aa than those of TaTGW-7Ab during grain development. High frequencies of the superior allele TaTGW-7Aa for higher TGW in Chinese mini-core collections (65.0%) and 501 wheat varieties (86.0%) indicated a strong and positive selection of this allele in wheat breeding. The molecular markers TGW7A and TG9 can be used for improvement of TGW in breeding programs. [ABSTRACT FROM AUTHOR]

The allelic variations of Vp-1B have been confirmed to have close association with seed dormancy (SD) and pre-harvest sprouting (PHS) of Chinese wheat in previous research, but little was known regarding whether the alleles of two other orthologs of Vp1 on 3AL ( Vp-1A) and 3DL ( Vp-1D) are also present and related to these traits. In view of this, 11 primer pairs flanking the whole sequences of these two orthologs were designed to investigate their allelic variations. The results identified six alleles of Vp-1A using the primer pair A17-19 among 81 wheat cultivars and advanced lines, which were designated as Vp-1Aa, Vp-1Ab, Vp-1Ac, Vp-1Ad, Vp-1Ae, and Vp-1Af. Except for Vp-1Ac, the other five alleles were proven novel, but no allelic variation was found in Vp-1D. On sequence analysis of alleles of Vp-1A, five deletions were observed, all occurring in the same region holding many TTC repeats. Of the six alleles detected in this study, four ( Vp-1Aa, Vp-1Ac, Vp-1Ae, and Vp-1Af) were generally distributed in varieties exhibiting higher average germination index (GI, range 0.46-0.56) and spike sprouting (SS, range 39.6-49.4%); however, the alleles Vp-1Ab and Vp-1Ad were distributed in genotypes carrying higher SD (GI 0.19-0.26) and stronger PHS resistance (SS 12.3-17.2%). On Spearman correlation analysis, the allele Vp-1Ab had significantly negative correlation with GI (−0.479) and SS (−0.542) at the 0.01 level, and the three alleles Vp-1Aa, Vp-1Ac, and Vp-1Ae had significantly positive correlation with GI [0.311 (0.05 level), 0.401 (0.01 level), and 0.294 (0.05 level)] and SS [0.283 (0.05 level), 0.309 (0.05 level), and 0.266 (0.05 level)]. The other alleles, including Vp-1Ad and Vp-1Af, also exhibited correlation, albeit not significant, with these two traits. This negative correlation showed that Vp-1Ab helped to improve SD and PHS tolerance, but Vp-1Aa, Vp-1Ac, and Vp-1Ae appeared to exert the opposite effect. To further confirm the association between alleles of Vp-1A and the two traits, a recombinant inbred line (RIL) population with 157 lines was genotyped using the primer pair A17-19, developed from the cross between Wanxianbaimaizi ( Vp-1Ab) and Jing411 ( Vp-1Ac). General linear model analysis indicated that variation in Vp-1A had a significant ( P < 0.001) association with the two traits, explaining 23.4% of the variation in GI and 16.7% of the variation in SS in the population across three crop seasons. [ABSTRACT FROM AUTHOR]

Tau normally associates with and stabilizes microtubules (MTs), but is hyperphosphorylated and aggregated into neurofibrillary tangles in Alzheimer's disease and related neurodegenerative diseases, which are collectively known as tauopathies. MTs are regulated by different forms of post-translational modification, including acetylation; acetylated MTs represent a more stable microtubule population. In our previous study, we showed that inhibition of histone deacetylase 6 (HDAC6), which deacetylates tubulin at lysine 40, rescues defects in MTs and in neuromuscular junction growth caused by tau overexpression. However, HDAC6 also acts on other proteins that are involved in distinct biological processes unrelated to tubulins. In order to examine directly the role of increased tubulin acetylation against tau toxicity, we generated a site-directed α-tubulinK40Q mutation by CRISPR/Cas9 technology to mimic the acetylated MTs and found that acetylation-mimicking α-tubulin rescued tau-induced MT defects and neuromuscular junction developmental abnormalities. We also showed that late administration of ACY-1215 and tubastatin A, two potent and selective inhibitors of HDAC6, rescued the tau-induced MT defects after the abnormalities had already become apparent. Overall, our results indicate that increasing MT acetylation by either genetic manipulations or drugs might be used as potential strategies for intervention in tauopathies.