Multiomics comparative analysis of the maize large grain mutant tc19 identified pathways related to kernel development.

BACKGROUND: The mechanism of grain development in elite maize breeding lines has not been fully elucidated. Grain length, grain width and grain weight are key components of maize grain yield. Previously, using the Chinese elite maize breeding line Chang7-2 and its large grain mutant tc19, we characterized the grain size developmental difference between Chang7-2 and tc19 and performed transcriptomic analysis. RESULTS: In this paper, using Chang7-2 and tc19, we performed comparative transcriptomic, proteomic and metabolomic analyses at different grain development stages. Through proteomics analyses, we found 2884, 505 and 126 differentially expressed proteins (DEPs) at 14, 21 and 28 days after pollination, respectively. Through metabolomics analysis, we identified 51, 32 and 36 differentially accumulated metabolites (DAMs) at 14, 21 and 28 days after pollination, respectively. Through multiomics comparative analysis, we showed that the phenylpropanoid pathways are influenced at transcriptomic, proteomic and metabolomic levels in all the three grain developmental stages. CONCLUSION: We identified several genes in phenylpropanoid biosynthesis, which may be related to the large grain phenotype of tc19. In summary, our results provided new insights into maize grain development.

OsCOMT, encoding a caffeic acid O-methyltransferase in melatonin biosynthesis, increases rice grain yield through dual regulation of leaf senescence and vascular development.

Melatonin, a natural phytohormone in plants, plays multiple critical roles in plant growth and stress responses. Although melatonin biosynthesis-related genes have been suggested to possess diverse biological functions, their roles and functional mechanisms in regulating rice grain yield remain largely unexplored. Here, we uncovered the roles of a caffeic acid O-methyltransferase (OsCOMT) gene in mediating rice grain yield through dual regulation of leaf senescence and vascular development. In vitro and in vivo evidence revealed that OsCOMT is involved in melatonin biosynthesis. Transgenic assays suggested that OsCOMT significantly delays leaf senescence at the grain filling stage by inhibiting degradation of chlorophyll and chloroplast, which, in turn, improves photosynthesis efficiency. In addition, the number and size of vascular bundles in the culms and leaves were significantly increased in the OsCOMT-overexpressing plants, while decreased in the knockout plants, suggesting that OsCOMT plays a positive role in vascular development of rice. Further evidence indicated that OsCOMT-mediated vascular development might owe to the crosstalk between melatonin and cytokinin. More importantly, we found that OsCOMT is a positive regulator of grain yield, and overexpression of OsCOMT increase grain yield per plant even in a high-yield variety background, suggesting that OsCOMT can be used as an important target for enhancing rice yield. Our findings shed novel insights into melatonin-mediated leaf senescence and vascular development and provide a possible strategy for genetic improvement of rice grain yield.

Origin, evolution and functional characterization of the land plant glycoside hydrolase subfamily GH5_11.

Colonization of the land by plants was a critical event in the establishment of modern terrestrial ecosystems, and many characteristics of land plants originated during this process, including the emergence of rosette terminal cellulose-synthesizing complexes. Cellulases are non-homologous isofunctional enzymes, encoded by glycosyl hydrolase (GH) gene families. Although the plant GH5_11 gene subfamily is presumed to encode a cell-wall degrading enzyme, its evolutionary and functional characteristics remain unclear. In the present study, we report the evolution of the land plant GH5_11 subfamily, and the functions of its members in terms of cellulase activity, through comprehensive phylogenetic analyses and observation of Arabidopsis mutants. Phylogenetic and sequence similarity analyses reveal that the ancestor of land plants acquired the GH5_11 gene from fungi through a horizontal gene transfer (HGT) event. Subsequently, positive selection with massive gene duplication and loss events contributed to the evolution of this subfamily in land plants. In Arabidopsis and rice, expression of GH5_11 genes are regulated by multiple abiotic stresses, the duplicated genes showing different patterns of expression. The Arabidopsis mutants atgh5_11a and atgh5_11c display low levels of cellulase and endoglucanase activities, with correspondingly high levels of cellulose, implying that the encoded proteins may function as endoglucanases. However, atgh5_11a and atgh5_11c also display an enlarged rosette leaf phenotype, and atgh5_11c is late-flowering under short photoperiods. These observations suggest that plant GH5_11s possess more functions beyond being endonucleases. To summarize, we demonstrate that the ancestor of land plants has acquired GH5_11 gene through HGT, which extends the cellulose degradation complexity. Our investigations illuminate features of part of the molecular framework underlying the origin of land plants and provide a focus on the cellulose degradation pathway.

Ancient horizontal transfer of transaldolase-like protein gene and its role in plant vascular development.

A major event in land plant evolution is the origin of vascular tissues, which ensure the long-distance transport of water, nutrients and organic compounds. However, the molecular basis for the origin and evolution of plant vascular tissues remains largely unknown. Here, we investigate the evolution of the land plant TAL-type transaldolase (TAL) gene and its potential function in rice (Oryza sativa) based on phylogenetic analyses and transgenic experiments, respectively. TAL genes are only present in land plants and bacteria. Phylogenetic analyses suggest that land plant TAL genes are derived from Actinobacteria through an ancient horizontal gene transfer (HGT) event. Further evidence reveals that land plant TAL genes have undergone positive selection and gained several introns following its acquisition by the most recent common ancestor of land plants. Transgenic plant experiments show that rice TAL is specifically expressed in vascular tissues and that knockdown of TAL expression leads to changes in both the number and pattern of vascular bundles. Our findings show that the ancient HGT of TAL from bacteria probably plays an important role in plant vascular development and adaptation to land environments.

Adaptive evolution and divergent expression of heat stress transcription factors in grasses.

BACKGROUND: Heat stress transcription factors (Hsfs) regulate gene expression in response to heat and many other environmental stresses in plants. Understanding the adaptive evolution of Hsf genes in the grass family will provide potentially useful information for the genetic improvement of modern crops to handle increasing global temperatures. RESULTS: In this work, we performed a genome-wide survey of Hsf genes in 5 grass species, including rice, maize, sorghum, Setaria, and Brachypodium, by describing their phylogenetic relationships, adaptive evolution, and expression patterns under abiotic stresses. The Hsf genes in grasses were divided into 24 orthologous gene clusters (OGCs) based on phylogeneitc relationship and synteny, suggesting that 24 Hsf genes were present in the ancestral grass genome. However, 9 duplication and 4 gene-loss events were identified in the tested genomes. A maximum-likelihood analysis revealed the effects of positive selection in the evolution of 11 OGCs and suggested that OGCs with duplicated or lost genes were more readily influenced by positive selection than other OGCs. Further investigation revealed that positive selection acted on only one of the duplicated genes in 8 of 9 paralogous pairs, suggesting that neofunctionalization contributed to the evolution of these duplicated pairs. We also investigated the expression patterns of rice and maize Hsf genes under heat, salt, drought, and cold stresses. The results revealed divergent expression patterns between the duplicated genes. CONCLUSIONS: This study demonstrates that neofunctionalization by changes in expression pattern and function following gene duplication has been an important factor in the maintenance and divergence of grass Hsf genes.

Analyses of sequence polymorphism and haplotype diversity of LEAFY genes revealed post-domestication selection in the Chinese elite maize inbred lines.

Post-domestication selection refers to the artificial selection on the loci controlling important agronomic traits during the process of genetic improvement in a population. The maize genes Zfl1 and Zfl2, duplicate orthologs of Arabidopsis LEAFY, are key regulators in plant branching, inflorescence and flower development, and reproduction. In this study, the full gene sequences of Zfl1 and Zfl2 from 62 Chinese elite inbred lines were amplified to evaluate their nucleotide polymorphisms and haplotype diversities. A total of 254 and 192 variants that included SNPs and indels were identified from the full sequences of Zfl1 and Zfl2, respectively. Although most of the variants were found to be located in the non-coding regions, the polymorphisms of CDS sequences classified Zfl1 into 16 haplotypes encoding 16 different proteins and Zfl2 into 18 haplotypes encoding eight different proteins. The population of Huangzaosi and its derived lines showed statistically significant signals of post-domestication selection on the Zfl1 CDS sequences, as well as lower nucleotide polymorphism and haplotype diversity than the whole set. However, the Zfl2 locus was only selected for in the heterotic group Reid. Further evidence revealed that at least 17 recombination events contributed to the genetic and haplotype diversities at the Zfl1 locus and 16 recombination events at the Zfl2 locus.

Evolution of land plant genes encoding L-Ala-D/L-Glu epimerases (AEEs) via horizontal gene transfer and positive selection.

BACKGROUND: The L-Ala-D/L-Glu epimerases (AEEs), a subgroup of the enolase superfamily, catalyze the epimerization of L-Ala-D/L-Glu and other dipeptides in bacteria and contribute to the metabolism of the murein peptide of peptidoglycan. Although lacking in peptidoglycan, land plants possess AEE genes that show high similarity to those in bacteria. RESULTS: Similarity searches revealed that the AEE gene is ubiquitous in land plants, from bryophytas to angiosperms. However, other eukaryotes, including green and red algae, do not contain genes encoding proteins with an L-Ala-D/L-Glu_epimerase domain. Homologs of land plant AEE genes were found to only be present in prokaryotes, especially in bacteria. Phylogenetic analysis revealed that the land plant AEE genes formed a monophyletic group with some bacterial homologs. In addition, land plant AEE proteins showed the highest similarity with these bacterial homologs and shared motifs only conserved in land plant and these bacterial AEEs. Integrated information on the taxonomic distribution, phylogenetic relationships and sequence similarity of the AEE proteins revealed that the land plant AEE genes were acquired from bacteria through an ancient horizontal gene transfer (HGT) event. Further evidence revealed that land plant AEE genes had undergone positive selection and formed the main characteristics of exon/intron structures through gaining some introns during the initially evolutionary period in the ancestor of land plants. CONCLUSIONS: The results of this study clearly demonstrated that the ancestor of land plants acquired an AEE gene from bacteria via an ancient HGT event. Other findings illustrated that adaptive evolution through positive selection has contributed to the functional adaptation and fixation of this gene in land plants.

Metabolomics reveals the response of hydroprimed maize to mitigate the impact of soil salinization.

Soil salinization is a major environmental stressor hindering global crop production. Hydropriming has emerged as a promising approach to reduce salt stress and enhance crop yields on salinized land. However, a better mechanisitic understanding is required to improve salt stress tolerance. We used a biochemical and metabolomics approach to study the effect of salt stress of hydroprimed maize to identify the types and variation of differentially accumulated metabolites. Here we show that hydropriming significantly increased catalase (CAT) activity, soluble sugar and proline content, decreased superoxide dismutase (SOD) activity and peroxide (H2O2) content. Conversely, hydropriming had no significant effect on POD activity, soluble protein and MDA content under salt stress. The Metabolite analysis indicated that salt stress significantly increased the content of 1278 metabolites and decreased the content of 1044 metabolites. Ethisterone (progesterone) was the most important metabolite produced in the roots of unprimed samples in response to salt s tress. Pathway enrichment analysis indicated that flavone and flavonol biosynthesis, which relate to scavenging reactive oxygen species (ROS), was the most significant metabolic pathway related to salt stress. Hydropriming significantly increased the content of 873 metabolites and significantly decreased the content of 1313 metabolites. 5-Methyltetrahydrofolate, a methyl donor for methionine, was the most important metabolite produced in the roots of hydroprimed samples in response to salt stress. Plant growth regulator, such as melatonin, gibberellin A8, estrone, abscisic acid and brassinolide involved in both treatment. Our results not only verify the roles of key metabolites in resisting salt stress, but also further evidence that flavone and flavonol biosynthesis and plant growth regulator relate to salt tolerance.

The plant streptolysin S (SLS)-associated gene B confers nitroaromatic tolerance and detoxification.

Nitroaromatic compounds, as the important chemical feedstock, have caused widespread environmental contaminations, and exhibited high toxicity and mutagenic activity to nearly all living organisms. The clean-up of nitroaromatic-contaminated soil and water has long been a major international concern. Here, we uncovered the role of a novel nitroreductase family gene, streptolysin S (SLS)-associated gene B (SagB), in enhancing nitroaromatic tolerance and detoxification of plants, and its potential application in phytoremediation of nitroaromatic contaminations. The expression of both the Arabidopsis and rice SagB genes is significantly induced by multiple hazardous nitroaromatic substances, including explosive pollutant 2,4,6-trinitrotoluene (TNT), natural compound 1-nitropyrene (1-NP) and herbicide pendimethalin (Pen). In vitro and in vivo evidences revealed that plant SagBs possess activities in degradation of these nitroaromatic substances. Arabidopsis and rice transgenic assays suggested that plant SagB genes increase tolerance and detoxification of nitroaromatic through facilitating its transformation to the amino derivative. More importantly, overexpression of plant SagBs increase their ability in TNT uptake, and remove more TNT from the growth culture. Our findings shed novel insights into a plant endogenous nitroreductase-mediated nitroaromatic tolerance and detoxification, and provide a new gene target for phytoremediation of nitroaromatic-contaminated environments.

Global Identification and Systematic Analysis of Lysine Malonylation in Maize (Zea mays L.).

Lysine malonylation is a kind of post-translational modifications (PTMs) discovered in recent years, which plays an important regulatory role in plants. Maize (Zea mays L.) is a major global cereal crop. Immunoblotting revealed that maize was rich in malonylated proteins. We therefore performed a qualitative malonylome analysis to globally identify malonylated proteins in maize. In total, 1,722 uniquely malonylated lysine residues were obtained in 810 proteins. The modified proteins were involved in various biological processes such as photosynthesis, ribosome and oxidative phosphorylation. Notably, a large proportion of the modified proteins (45%) were located in chloroplast. Further functional analysis revealed that 30 proteins in photosynthesis and 15 key enzymes in the Calvin cycle were malonylated, suggesting an indispensable regulatory role of malonylation in photosynthesis and carbon fixation. This work represents the first comprehensive survey of malonylome in maize and provides an important resource for exploring the function of lysine malonylation in physiological regulation of maize.

Nucleotide Diversity of the Maize ZmCNR13 Gene and Association With Ear Traits.

The maize (Zea mays L.) ZmCNR13 gene, encoding a protein of fw2.2-like (FWL) family, has been demonstrated to be involved in cell division, expansion, and differentiation. In the present study, the genomic sequences of the ZmCNR13 locus were re-sequenced in 224 inbred lines, 56 landraces and 30 teosintes, and the nucleotide polymorphism and selection signature were estimated. A total of 501 variants, including 415 SNPs and 86 Indels, were detected. Among them, 51 SNPs and 4 Indels were located in the coding regions. Although neutrality tests revealed that this locus had escaped from artificial selection during the process of maize domestication, the population of inbred lines possesses lower nucleotide diversity and decay of linkage disequilibrium. To estimate the association between sequence variants of ZmCNR13 and maize ear characteristics, a total of ten ear-related traits were obtained from the selected inbred lines. Four variants were found to be significantly associated with six ear-related traits. Among them, SNP2305, a non-synonymous mutation in exon 2, was found to be associated with ear weight, ear grain weight, ear diameter and ear row number, and explained 4.59, 4.61, 4.31, and 8.42% of the phenotypic variations, respectively. These results revealed that natural variations of ZmCNR13 might be involved in ear development and can be used in genetic improvement of maize ear-related traits.

Combined bulked segregant sequencing and traditional linkage analysis for identification of candidate gene for purple leaf sheath in maize.

Anthocyanin accumulation in various maize tissues plays important roles in plant growth and development. In addition, some color-related traits can be used as morphological markers in conventional maize breeding processes and purity identification of hybrid seeds. Here, we noticed that the leaf sheath color was controlled by a dominant gene, because purple (PSH) and green leaf sheaths (GSH) were separated at a ratio of 3:1 in an F2 population. To map the gene, an F2 and a recombinant inbred line (RIL) population were derived from a cross between inbred line T877 (PSH) and DH1M (GSH). The PSH locus was mapped to the genomic region within 128.8 to 138.4 Mb using a bulked segregant sequencing approach. This position was further validated by linkage mapping using 190 F2 plants with GSH. Subsequently, the PSH locus was fine-mapped into an interval of 304.2 kb. A maize gene, GRMZM5G822829, was identified in this region, encoding a bHLH transcription factor. The expression level of this gene in T877 was found to be 9-fold higher than that of DH1M. In conclusion, our results suggest that GRMZM5G822829 is the putative candidate gene conferring leaf sheath color in maize.

Correction: Combined bulked segregant sequencing and traditional linkage analysis for identification of candidate gene for purple leaf sheath in maize.

[This corrects the article DOI: 10.1371/journal.pone.0190670.].

Ancestor of land plants acquired the DNA-3-methyladenine glycosylase (MAG) gene from bacteria through horizontal gene transfer.

The origin and evolution of land plants was an important event in the history of life and initiated the establishment of modern terrestrial ecosystems. From water to terrestrial environments, plants needed to overcome the enhanced ultraviolet (UV) radiation and many other DNA-damaging agents. Evolving new genes with the function of DNA repair is critical for the origin and radiation of land plants. In bacteria, the DNA-3-methyladenine glycosylase (MAG) recognizes of a variety of base lesions and initiates the process of the base excision repair for damaged DNA. The homologs of MAG gene are present in all major lineages of streptophytes, and both the phylogenic and sequence similarity analyses revealed that green plant MAG gene originated through an ancient horizontal gene transfer (HGT) event from bacteria. Experimental evidence demonstrated that the expression of the maize ZmMAG gene was induced by UV and zeocin, both of which are known as DNA-damaging agents. Further investigation revealed that Streptophyta MAG genes had undergone positive selection during the initial evolutionary period in the ancestor of land plants. Our findings demonstrated that the ancient HGT of MAG to the ancestor of land plants probably played an important role in preadaptation to DNA-damaging agents in terrestrial environments.

Meta-Analysis of the SLCO1B1 c.521T>C Variant Reveals Slight Influence on the Lipid-Lowering Efficacy of Statins.

BACKGROUND: Several studies have focused on the association between the lipid-lowering efficacy of statins and the SLCO1B1 c.521T>C polymorphism; however, the results are conflicting. The effects of statins show significant variability between individuals. This meta-analysis aimed to investigate the effects of the SLCO1B1 c.521T>C polymorphism on the lipid-lowering effects of statins. METHODS: We systematically searched PubMed and Web of Science to screen relevant studies. Meta-analysis was performed to identify the association between SLCO1B1 c.521 polymorphisms and the lipid-lowering effects of statins on the basis of the standard mean difference (SMD) and 95% confidence intervals (CIs). Additionally, we checked for heterogeneity (I(2)) among studies and evidence of publication bias. We obtained eight studies including 2,012 wild genotype (T/T) and 526 variant genotype (T/C and C/C) cases. RESULTS: No significant difference was observed in the lipid-lowering efficacy of statins between the wild and variant genotypes of SLCO1B1, with a pooled SMD of 0.03 (95% CI: -0.07-0.13). Furthermore, there was no significant effect in the meta-analyses of the variant heterozygote, homozygote, and Chinese populations. Subgroup meta-analysis indicated that the time required for the statin to take effect did not significantly affect the association between lipid-lowering efficacy of statins and SLCO1B1 c.521T>C polymorphism. However, the wild genotype improved the lipid-lowering efficacy of simvastatin with a pooled SMD of -0.26 (95% CI: -0.47- -0.05). CONCLUSIONS: No significant association was detected between the lipid-lowering efficacy of statins and the SLCO1B1 c.521T>C polymorphism, with the exception of simvastatin.

Non-invasive immunodiagnosis of Schistosomiasis japonica: the detection of specific antibodies in saliva.

OBJECTIVE: To assess the feasibility of using saliva for Schistosomiasis japonica diagnosis. METHODS: Schistosoma japonicum infected animal model was established. Pairs of saliva and serum samples from rabbits and chronic schistosomiasis patients were collected. Anti-schistosoma specific antibodies in saliva and serum were detected by indirect ELISA. RESULTS: The specificities of antibody detection of rabbit saliva and serum were 93% (28/30) and 97% (29/30), respectively, and the sensitivities of antibody detection of rabbit serum and saliva were 100% (24/24) and 88% (21/24), respectively. A significant correlation (r = 0.5307, P = 0.0038 < 0.05) existed between anti-SEA IgG levels in serum and saliva. As with those in serum, anti-SEA IgG levels in saliva could reflect the state of infection and treatment. The sensitivity of antibody detection was 91% (29/32) for patient saliva samples and 100% (32/32) for their sera. 8 samples were positive in 140 normal saliva samples (i.e. 6% false positive rate) and 6 samples were positive in 156 normal serum samples (4% false positive rate). There was a significant correlation (r = 0.4227, P = 0.008 < 0.05) between specific antibodies in saliva and serum. CONCLUSION: The detection of specific antibodies in saliva can be used as a non-invasive immunodiagnosis method of Schistosomiasis japonica.

Nucleotide diversity of Maize ZmBT1 gene and association with starch physicochemical properties.

Cereal Brittle1 protein has been demonstrated to be involved in the ADP-Glc transport into endosperm plastids, and plays vital roles in the biosynthesis of starch. In this study, the genomic sequences of the ZmBT1 gene in 80 elite maize inbred lines were obtained, and the nucleotide polymorphisms and haplotype diversity were detected. A total of 30 variants, including 22 SNPs and 8 indels, were detected from the full sequences of this gene. Among these polymorphic sites, 9 SNPs and 2 indels were found to be located in the coding region. The polymorphisms of CDS sequences classified the maize ZmBT1 gene into 6 haplotypes, which encode 6 different ZmBT1 proteins. Neutrality tests revealed a decrease in population size and/or balancing selection on the maize ZmBT1 locus. To detect the association between sequence variations of this gene and the starch physicochemical properties, 7 pasting and 4 gelatinization traits of starch were measured for the tested inbred lines using rapid visco analyzer (RVA) and differential scanning calorimeter (DSC), respectively. The result of association analysis revealed that an indel in the coding region was significantly associated with the phenotypic variation of starch gelatinization enthalpy.

Nucleotide polymorphisms and haplotype diversity of RTCS gene in China elite maize inbred lines.

The maize RTCS gene, encoding a LOB domain transcription factor, plays important roles in the initiation of embryonic seminal and postembryonic shoot-borne root. In this study, the genomic sequences of this gene in 73 China elite inbred lines, including 63 lines from 5 temperate heteroric groups and 10 tropic germplasms, were obtained, and the nucleotide polymorphisms and haplotype diversity were detected. A total of 63 sequence variants, including 44 SNPs and 19 indels, were identified at this locus, and most of them were found to be located in the regions of UTR and intron. The coding region of this gene in all tested inbred lines carried 14 haplotypes, which encoding 7 deferring RTCS proteins. Analysis of the polymorphism sites revealed that at least 6 recombination events have occurred. Among all 6 groups tested, only the P heterotic group had a much lower nucleotide diversity than the whole set, and selection analysis also revealed that only this group was under strong negative selection. However, the set of Huangzaosi and its derived lines possessed a higher nucleotide diversity than the whole set, and no selection signal were identified.