Male Sterile Wheat Research Articles

Identification of Proteins Involved in Carbohydrate Metabolism and Energy Metabolism Pathways and Their Regulation of Cytoplasmic Male Sterility in Wheat.

Cytoplasmic male sterility (CMS) where no functional pollen is produced has important roles in wheat breeding. The anther is a unique organ for male gametogenesis and its abnormal development can cause male sterility. However, the mechanisms and regulatory networks related to plant male sterility are poorly understood. In this study, we conducted comparative analyses using isobaric tags for relative and absolute quantification (iTRAQ) of the pollen proteins in a CMS line and its wheat maintainer. Differentially abundant proteins (DAPs) were analyzed based on Gene Ontology classifications, metabolic pathways and transcriptional regulation networks using Blast2GO. We identified 5570 proteins based on 23,277 peptides, which matched with 73,688 spectra, including proteins in key pathways such as glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase and 6-phosphofructokinase 1 in the glycolysis pathway, isocitrate dehydrogenase and citrate synthase in the tricarboxylic acid cycle and nicotinamide adenine dinucleotide (NADH)-dehydrogenase and adenosine-triphosphate (ATP) synthases in the oxidative phosphorylation pathway. These proteins may comprise a network that regulates male sterility in wheat. Quantitative real time polymerase chain reaction (qRT-PCR) analysis, ATP assays and total sugar assays validated the iTRAQ results. These DAPs could be associated with abnormal pollen grain formation and male sterility. Our findings provide insights into the molecular mechanism related to male sterility in wheat.

Mitochondrial Dysfunction Causes Oxidative Stress and Tapetal Apoptosis in Chemical Hybridization Reagent-Induced Male Sterility in Wheat.

Male sterility in plants has been strongly linked to mitochondrial dysfunction. Chemical hybridization agent (CHA)-induced male sterility is an important tool in crop heterosis. Therefore, it is important to better understand the relationship between mitochondria and CHA-induced male sterility in wheat. This study reports on the impairment of mitochondrial function duo to CHA-SQ-1, which occurs by decreasing cytochrome oxidase and adenosine triphosphate synthase protein levels and theirs activities, respiratory rate, and in turn results in the inhibition of the mitochondrial electron transport chain (ETC), excessive production of reactive oxygen species (ROS) and disruption of the alternative oxidase pathway. Subsequently, excessive ROS combined with MnSOD defects results in damage to the mitochondrial membrane, followed by ROS release into the cytoplasm. The microspores underwent severe oxidative stress during pollen development. Furthermore, chronic oxidative stress, together with the overexpression of type II metacaspase, triggered premature tapetal apoptosis, which resulted in pollen abortion. Accordingly, we propose a metabolic pathway for mitochondrial-mediated male sterility in wheat, which provides information on the molecular events underlying CHA-SQ-1-induced abortion of anthers and may serve as an additional guide to the practical application of hybrid breeding.

Chemical hybridizing agent SQ-1-induced male sterility in Triticum aestivum L.: a comparative analysis of the anther proteome

BackgroundHeterosis is widely used to increase the yield of many crops. However, as wheat is a self-pollinating crop, hybrid breeding is not so successful in this organism. Even though male sterility induced by chemical hybridizing agents is an important aspect of crossbreeding, the mechanisms by which these agents induce male sterility in wheat is not well understood.ResultsWe performed proteomic analyses using the wheat Triticum aestivum L.to identify those proteins involved in physiological male sterility (PHYMS) induced by the chemical hybridizing agent CHA SQ-1. A total of 103 differentially expressed proteins were found by 2D–PAGE and subsequently identified by MALDI-TOF/TOF MS/MS. In general, these proteins had obvious functional tendencies implicated in carbohydrate metabolism, oxidative stress and resistance, protein metabolism, photosynthesis, and cytoskeleton and cell structure. In combination with phenotypic, tissue section, and bioinformatics analyses, the identified differentially expressed proteins revealed a complex network behind the regulation of PHYMS and pollen development. Accordingly, we constructed a protein network of male sterility in wheat, drawing relationships between the 103 differentially expressed proteins and their annotated biological pathways. To further validate our proposed protein network, we determined relevant physiological values and performed real-time PCR assays.ConclusionsOur proteomics based approach has enabled us to identify certain tendencies in PHYMS anthers. Anomalies in carbohydrate metabolism and oxidative stress, together with premature tapetum degradation, may be the cause behind carbohydrate starvation and male sterility in CHA SQ-1 treated plants. Here, we provide important insight into the mechanisms underlying CHA SQ-1-induced male sterility. Our findings have practical implications for the application of hybrid breeding in wheat.

Phenotype characterisation and analysis of expression patterns of genes related mainly to carbohydrate metabolism and sporopollenin in male-sterile anthers induced by high temperature in wheat (Triticum aestivum)

Male reproductive development in higher plants is highly sensitive to various stressors, including high temperature (HT). In this study, physiological male-sterile plants of wheat (Triticum aestivum L.) were established using HT induction. The physiological changes and expression levels of genes mainly related to carbohydrate metabolism and sporopollenin in male-sterile processes were studied by using biological techniques, including iodine–potassium iodide staining, paraffin sectioning, scanning electron microscopy (SEM) and fluorescent quantitative analysis. Results of paraffin sectioning and SEM revealed that parts of HT male-sterile anthers, including the epidermis and tapetum, were remarkably different from those of normal anthers. The expression levels of TaSUT1, TaSUT2, IVR1 and IVR5 were significantly lower than of normal anthers at the early microspore and trinucleate stages. The RAFTIN1 and TaMS26 genes may contribute to biosynthesis and proper ‘fixation’ of sporopollenin in the development of pollen wall; however, their expression levels were significantly higher at the early tetrad stage and early microspore stage in HT sterile anthers. The recently cloned MS1 gene was expressed at the early tetrad and early microspore stages but not at the trinucleate stage. Moreover, this gene showed extremely significant, high expression in HT sterile anthers compared with normal anthers. These results demonstrate that HT induction of wheat male sterility is probably related to the expression of genes related to carbohydrate metabolism and sporopollenin metabolism. This provides a theoretical basis and technological approach for further studies on the mechanisms of HT induction of male sterility.

Uncovering Male Fertility Transition Responsive miRNA in a Wheat Photo-Thermosensitive Genic Male Sterile Line by Deep Sequencing and Degradome Analysis.

MicroRNAs (miRNAs) are endogenous small RNAs which play important negative regulatory roles at both the transcriptional and post-transcriptional levels in plants. Wheat is the most commonly cultivated plant species worldwide. In this study, RNA-seq analysis was used to examine the expression profiles of miRNA in the spikelets of photo-thermosenisitive genic male sterile (PTGMS) wheat line BS366 during male fertility transition. Through mapping on their corresponding precursors, 917–7,762 novel miRNAs were found in six libraries. Six novel miRNAs were selected for examination of their secondary structures and confirmation by stem-loop RT-PCR. In a differential expression analysis, 20, 22, and 58 known miRNAs exhibited significant differential expression between developmental stages 1 (secondary sporogenous cells had formed), 2 (all cells layers were present and mitosis had ceased), and 3 (meiotic division stage), respectively, of fertile and sterile plants. Some of these differential expressed miRNAs, such as tae-miR156, tae-miR164, tae-miR171, and tae-miR172, were shown to be associated with their targets. These targets were previously reported to be related to pollen development and/or male sterility, indicating that these miRNAs and their targets may be involved in the regulation of male fertility transition in the PTGMS wheat line BS366. Furthermore, target genes of miRNA cleavage sites were validated by degradome sequencing. In this study, a possible signal model for the miRNA-mediated signaling pathway during the process of male fertility transition in the PTGMS wheat line BS366 was developed. This study provides a new perspective for understanding the roles of miRNAs in male fertility in PTGMS lines of wheat.

Proteomics Analysis of Photo-Thermo-Sensitive Male Sterility Wheat Line BNS during Its Thermosensitive Period

Photo-thermo sensitive male sterile (PTMS) line is one of the important materials in utilizing heterosis in crops. Wheat line BNS (Bainong sterility) is an important nuclear-controlling PTMS line and suitable for growing and seed production in Huang Huai wheat zone in China. It has genetic stability with male sterility when sowing in autumn and male fertility when sowing in spring. Their thermosensitive periods were between stamen and pistil differentiation stage and anther connective stage and they could be regarded key periods for fertility conversion in BNS. To determine the molecular mechanisms of fertility conversion at thermosensitive period, we investigated characters of seed setting, anther and pollen grain of fertile and sterile BNS plants and compared young spike proteome patterns at their thermosensitive periods between the two BNS plants. The results showed that sterile plants had lower seed setting rate and pollen number, small pollen grain and lower pollen vitality than fertile plants. Out of protein spots reproducibly detected and analyzed on two dimensional electrophoresis gels, 76 spots showed significant changes in at least one BNS plant and 36 spots were identified by MALDI-TOF MS. The results showed that proteins involved in multiple biochemical pathways were differentially expressed at thermosensitive period between the two plants, including energy metabolism, stress response, signal transduction and regulation, protein process, amino acid and fatty acid metabolism, nucleic acid metabolism etc. Some of these proteins are reported to be involved in the abortion of anther or pollen grains in MS plants, such as energy metabolism and anti oxidative stress, and some were found to be novel proteins involved in the fertility conversion, such as phytohormones regulation. These results indicated that proteins related with anther or pollen development had expressed differently between the two BNS plants before anther development and phytohormones and signal transduction might be involved in the regulation of fertility conversion at thermosensitive period. Our studies have provided new insight to reveal the molecular mechanisms of fertility conversion at thermosensitive period in PTMS wheat.

A TRIM insertion in the promoter of Ms2 causes male sterility in wheat

The male-sterile ms2 mutant has been known for 40 years and has become extremely important in the commercial production of wheat. However, the gene responsible for this phenotype has remained unknown. Here we report the map-based cloning of the Ms2 gene. The Ms2 locus is remarkable in several ways that have implications in basic biology. Beyond having no functional annotation, barely detectable transcription in fertile wild-type wheat plants, and accumulated destructive mutations in Ms2 orthologs, the Ms2 allele in the ms2 mutant has acquired a terminal-repeat retrotransposon in miniature (TRIM) element in its promoter. This TRIM element is responsible for the anther-specific Ms2 activation that confers male sterility. The identification of Ms2 not only unravels the genetic basis of a historically important breeding trait, but also shows an example of how a TRIM element insertion near a gene can contribute to genetic novelty and phenotypic plasticity.

Wheat Ms2 encodes for an orphan protein that confers male sterility in grass species

Male sterility is a valuable trait for plant breeding and hybrid seed production. The dominant male-sterile gene Ms2 in common wheat has facilitated the release of hundreds of breeding lines and cultivars in China. Here, we describe the map-based cloning of the Ms2 gene and show that Ms2 confers male sterility in wheat, barley and Brachypodium. MS2 appears as an orphan gene within the Triticinae and expression of Ms2 in anthers is associated with insertion of a retroelement into the promoter. The cloning of Ms2 has substantial potential to assemble practical pipelines for recurrent selection and hybrid seed production in wheat.

Genome-wide characterization of JASMONATE-ZIM DOMAIN transcription repressors in wheat (Triticum aestivum L.)

BackgroundThe JASMONATE-ZIM DOMAIN (JAZ) repressor family proteins are jasmonate co-receptors and transcriptional repressor in jasmonic acid (JA) signaling pathway, and they play important roles in regulating the growth and development of plants. Recently, more and more researches on JAZ gene family are reported in many plants. Although the genome sequencing of common wheat (Triticum aestivum L.) and its relatives is complete, our knowledge about this gene family remains vacant.ResultsFourteen JAZ genes were identified in the wheat genome. Structural analysis revealed that the TaJAZ proteins in wheat were as conserved as those in other plants, but had structural characteristics. By phylogenetic analysis, all JAZ proteins from wheat and other plants were clustered into 11 sub-groups (G1-G11), and TaJAZ proteins shared a high degree of similarity with some JAZ proteins from Aegliops tauschii, Brachypodium distachyon and Oryza sativa. The Ka/Ks ratios of TaJAZ genes ranged from 0.0016 to 0.6973, suggesting that the TaJAZ family had undergone purifying selection in wheat. Gene expression patterns obtained by quantitative real-time PCR (qRT-PCR) revealed differential temporal and spatial regulation of TaJAZ genes under multifarious abiotic stress treatments of high salinity, drought, cold and phytohormone. Among these, TaJAZ7, 8 and 12 were specifically expressed in the anther tissues of the thermosensitive genic male sterile (TGMS) wheat line BS366 and normal control wheat line Jing411. Compared with the gene expression patterns in the normal wheat line Jing411, TaJAZ7, 8 and 12 had different expression patterns in abnormally dehiscent anthers of BS366 at the heading stage 6, suggesting that specific up- or down-regulation of these genes might be associated with the abnormal anther dehiscence in TGMS wheat line.ConclusionThis study analyzed the size and composition of the JAZ gene family in wheat, and investigated stress responsive and differential tissue-specific expression profiles of each TaJAZ gene in TGMS wheat line BS366. In addition, we isolated 3 TaJAZ genes that would be more likely to be involved in the regulation of abnormal anther dehiscence in TGMS wheat line. In conclusion, the results of this study contributed some novel and detailed information about JAZ gene family in wheat, and also provided 3 potential candidate genes for improving the TGMS wheat line.

Efficacy of chemical hybridizing agents for inducing male sterility in wheat

The present investigations were carried out to induce male sterility in wheat by using different chemical hybridizing agents (CHAs) at CCS Haryana Agricultural University, Hisar, Haryana. The genotypes exhibited differential response to CHAs in relation to induction of male sterility at similar doses and time of spray. Higher doses of all chemicals induced more male sterility than the lower doses in all the genotypes included in the study. CHC series of chemicals viz CHC 9901, CHC 9902, CHC 9903, CHC 9904 and CHC 9905 were found effective in inducing high level of male sterility(>90%). Among CH series CH 0288 at 2000 ppm induced maximum male sterility(82.42%)in genotype UP 2425. The genotype UP 2425 was found most responsive in relation to male sterility induction followed by PBW 343, WH 542 and UP 2338. But at the same time it was also found that these genotypes were affected with female sterility. Genotype PBW 343 was observed most sensitive to CHA treatments with respect to male sterility. Seed setting was also affected by female sterility.

DNA methylation of the TAA1 gene regulates formation of the pollen wall in chemically induced male sterility in wheat (Triticum aestivum)

DNA methylation is an important epigenetic modification that may contribute to environmentally induced phenotypic variations by regulating gene expression. Chemically induced male sterility (CIMS) lines in wheat (Triticum aestivum L.) can transform from sterile to fertile, induced by a chemical hybridising agent during anther development. So far, little is known about the DNA methylation variation of CIMS in wheat. TAA1 regulates pollen wall development, probably through converting fatty acids to fatty alcohol in wheat. We investigated the DNA methylation pattern of the TAA1 gene in the core promoter region by using the bisulfite genomic sequencing method, and higher methylation was observed in CIMS. The expression levels of the TAA1 gene were also evaluated by real time quantitative reverse transcriptase PCR analysis, which revealed that the expression levels of the TAA1 gene were downregulated in CIMS. The aliphatic composition of the anther underwent accumulation in line 1376-CIMS, revealed by gas chromatography–mass spectrometry, including increments of tetradecanoic acid, hexadecanoic acid and octadecanoic acid. Scanning electron microscopy revealed that anther and pollen wall formation was significantly altered in 1376-CIMS.These results suggested that DNA methylation of the TAA1 gene may be involved in the sterility–fertility transition of CIMS.

Comprehensive analyses of the annexin gene family in wheat.

BackgroundAnnexins are an evolutionarily conserved multigene family of calcium-dependent phospholipid binding proteins that play important roles in stress resistance and plant development. They have been relatively well characterized in model plants Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa), but nothing has been reported in hexaploid bread wheat (Triticum aestivum) and barely (Hordeum vulgare), which are the two most economically important plants.ResultsBased on available genomic and transcriptomic data, 25 and 11 putative annexin genes were found through in silico analysis in wheat and barley, respectively. Additionally, eight and 11 annexin genes were identified from the draft genome sequences of Triticum urartu and Aegilops tauschii, progenitor for the A and D genome of wheat, respectively. By phylogenetic analysis, annexins in these four species together with other monocots and eudicots were classified into six different orthologous groups. Pi values of each of Ann1–12 genes among T. aestivum, T. urartu, A. tauschii and H. vulgare species was very low, with the exception of Ann2 and Ann5 genes. Ann2 gene has been under positive selection, but Ann6 and Ann7 have been under purifying selection among the four species in their evolutionary histories. The nucleotide diversities of Ann1–12 genes in the four species were 0.52065, 0.59239, 0.60691 and 0.53421, respectively. No selective pressure was operated on annexin genes in the same species. Gene expression patterns obtained by real-time PCR and re-analyzing the public microarray data revealed differential temporal and spatial regulation of annexin genes in wheat under different abiotic stress conditions such as salinity, drought, cold and abscisic acid. Among those genes, TaAnn10 is specifically expressed in the anther but fails to be induced by low temperature in thermosensitive genic male sterile lines, suggesting that specific down-regulation of TaAnn10 is associated with conditional male sterility in wheat.ConclusionsThis study analyzed the size and composition of the annexin gene family in wheat and barley, and investigated differential tissue-specific and stress responsive expression profiles of the gene family in wheat. These results provided significant information for understanding the diverse roles of plant annexins and opened a new avenue for functional studies of cold induced male sterility in wheat.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2750-y) contains supplementary material, which is available to authorized users.

Investigating Pollen and Gene Flow of WYMV-Resistant Transgenic Wheat N12-1 Using a Dwarf Male-Sterile Line as the Pollen Receptor.

Pollen-mediated gene flow (PMGF) is the main mode of transgene flow in flowering plants. The study of pollen and gene flow of transgenic wheat can help to establish the corresponding strategy for preventing transgene escape and contamination between compatible genotypes in wheat. To investigate the pollen dispersal and gene flow frequency in various directions and distances around the pollen source and detect the association between frequency of transgene flow and pollen density from transgenic wheat, a concentric circle design was adopted to conduct a field experiment using transgenic wheat with resistance to wheat yellow mosaic virus (WYMV) as the pollen donor and dwarf male-sterile wheat as the pollen receptor. The results showed that the pollen and gene flow of transgenic wheat varied significantly among the different compass sectors. A higher pollen density and gene flow frequency was observed in the downwind SW and W sectors, with average frequencies of transgene flow of 26.37 and 23.69% respectively. The pollen and gene flow of transgenic wheat declined dramatically with increasing distance from its source. Most of the pollen grains concentrated within 5 m and only a few pollen grains were detected beyond 30 m. The percentage of transgene flow was the highest where adjacent to the pollen source, with an average of 48.24% for all eight compass directions at 0 m distance. Transgene flow was reduced to 50% and 95% between 1.61 to 3.15 m, and 10.71 to 20.93 m, respectively. Our results suggest that climate conditions, especially wind direction, may significantly affect pollen dispersal and gene flow of wheat. The isolation-by-distance model is one of the most effective methods for achieving stringent transgene confinement in wheat. The frequency of transgene flow is directly correlated with the relative density of GM pollen grains in air currents, and pollen competition may be a major factor influencing transgene flow.

Proteome analysis of pollen in the K-type male sterility line 732A and its maintainer 732B in wheat (Triticum aestivum L.) by two-dimensional gel electrophoresis

Male sterility, especially cytoplasmic male sterility (CMS) where no functional pollen is produced, plays an important role in wheat breeding. Some specific proteins are related to male sterility in wheat, especially those present in pollen. To elucidate these sterility-related proteins, we applied two-dimensional gel electrophoresis (2-DE) to the proteomic analysis of pollen from a CMS line (732A) and its maintainer (732B) in wheat (Triticum aestivum L.), where trichloroacetic acid/acetone precipitation was used to extract pollen proteins from the wheat anthers. Analysis using PDQuest detected >350 reproducible protein spots on each 2-DE gel, where pI 4–7. In total, 41 spots were found to be differentially expressed, where 7/41 were identified by MALDI TOF/TOF–mass spectrometry analysis and protein database searching. The proteins comprised NADPH-producing dehydrogenase in the oxidative pentose phosphate pathway, cinnamyl alcohol dehydrogenase, cytosolic 3-phosphoglycerate kinase, caffeic acid O-methyltransferase, ascorbate peroxidase, glutamine synthetase isoform GS1a, and one unknown protein. In the uninucleate stage, all of the identified protein spots were downregulated in 732A, except the unknown protein, whereas there were no obvious differences in 732B. In the binucleate stage, the expression level of glutamine synthetase isoform GS1a did not differ between 732A and 732B, but caffeic acid O-methyltransferase was downregulated in 732A and the other five proteins were upregulated in 732A. These results suggest that male sterility in 732A is related to energy metabolism perturbation, active oxygen accumulation, programmed cell death, the pentose phosphate pathway, and glycolysis.

TaOPR2 encodes a 12-oxo-phytodienoic acid reductase involved in the biosynthesis of jasmonic acid in wheat (Triticum aestivum L.)

The 12-oxo-phytodienoic acid reductases (OPRs) are involved in the various processes of growth and development in plants, and classified into the OPRⅠ and OPRⅡ subgroups. In higher plants, only OPRⅡ subgroup genes take part in the biosynthesis of endogenous jasmonic acid. In this study, we isolated a novel OPRⅡ subgroup gene named TaOPR2 (GeneBank accession: KM216389) from the thermo-sensitive genic male sterile (TGMS) wheat cultivar BS366. TaOPR2 was predicted to encode a protein with 390 amino acids. The encoded protein contained the typical oxidored_FMN domain, the C-terminus peroxisomal-targeting signal peptide, and conserved FMN-binding sites. TaOPR2 was mapped to wheat chromosome 7B and located on peroxisome. Protein evolution analysis revealed that TaOPR2 belongs to the OPRⅡ subgroup and shares a high degree of identity with other higher plant OPR proteins. The quantitative real-time PCR results indicated that the expression of TaOPR2 is inhibited by abscisic acid (ABA), salicylic acid (SA), gibberellic acid (GA3), low temperatures and high salinity. In contrast, the expression of TaOPR2 can be induced by wounding, drought and methyl jasmonate (MeJA). Furthermore, the transcription level of TaOPR2 increased after infection with Puccinia striiformis f. sp. tritici and Puccinia recondite f. sp. tritici. TaOPR2 has NADPH-dependent oxidoreductase activity. In addition, the constitutive expression of TaOPR2 can rescue the male sterility phenotype of Arabidopsis mutant opr3. These results suggest that TaOPR2 is involved in the biosynthesis of jasmonic acid (JA) in wheat.

Cytological characterization of a thermo-sensitive cytoplasmic male-sterile wheat line having K-type cytoplasm of Aegilops kotschyi.

Male sterility is an important tool for obtaining crop heterosis. A thermo-sensitive cytoplasmic male-sterile (TCMS) line was developed recently using a new method based on tiller regeneration. In the present study, we explored the critical growth stages required to maintain thermo-sensitive male sterility in TCMS lines and found that fertility is associated with abnormal tapetal and microspore development. We investigated the fertility and cytology of temperature-treated plant anthers at various developmental stages. TCMS line KTM3315A exhibited thermo-sensitive male sterility in Zadoks growth stages 41–49 and 58–59. Morphologically, the line exhibited thermo-sensitive male sterility at 3–9 days before heading and at 3–6 days before flowering, and it was partially restored in three locations during spring and summer. TCMS line KTM3315A plants exhibited premature tapetal programmed cell death (PCD) from the early uninucleate stage of microspore development until the tapetal cells degraded completely. Microspore development was then blocked and the pollen abortion type was stainable abortion. Thus, male fertility in the line KTM3315A is sensitive to temperature and premature tapetal PCD is the main cause of pollen abortion, where it determines the starting period and affects male fertility conversion in K-type TCMS lines at certain temperatures.

THE RELATIONSHIP BETWEEN MALE STERILITY AND MEMBRANE LIPID PEROXIDATION AND ANTIOXIDANT ENZYMES IN WHEAT (Triticum aestivum L.)

Malondialdehyde (MDA), which is thought to be important inducible factor of cell apoptosis if excessively accumulated in cells, is often regarded as a widely used marker of oxidative lipid injury whose concentration varies in response to biotic and abiotic stress. To elucidate the metabolic mechanism of ROS production and scavenging of the male sterility, normal fertility and male sterility wheat, including genetic and physiologically male sterility, by comparing MDA content and antioxidant enzymes changes in the sterile and fertile leaves, spikes, florets, rachillaes, ovaries and anthers at different developmental anther stages, were employed for studying the relationship between the male sterility and membrane lipid peroxidation, antioxidant enzymes. The results showed that tissue of male sterility had roughly higher content of MDA than corresponding fertile plants, and ovary performance was not obvious but most obvious in anther. Simultaneously, the higher activities of antioxidant enzymes were measured in the anthers of male sterility plant, in which an increase of ROS in anthers of abortion stage had an inducible effect on the antioxidant enzymes. MDA content and scavenging-enzyme activities had difference in the sterile and fertile leaves, spikes, florets, rachillaes at different anther stages, indicating that different tissues of ROS metabolism had noticeable difference in male sterility and normal wheat. It can be considered that the disorder of scavenging-enzyme activities and accumulation of MDA in the male organ, and supplier of nutrition blades were causing a further substance metabolism disorder, energy loss, leading to the occurrence of male sterility. Key words: Wheat - male sterility – malondialdehyde - antioxidant enzymes Abbreviations: CAT---Catalase; CMS---Cytoplasmic male sterility; CHA---Chemical Hybridization Agent; GS---Cytoplasmic male sterile line (S)-1376; MDA---Malondialdehyde; NW---Wheat cultivar 1376; PMS---CHA-SQ-1 induced male sterility wheat; POD---Peroxidase; ROS---Reactive oxygen species; SOD---Superoxide dismutase; SEM---Scanning electronm icroscopy.

The role of hydrogen peroxide in wheat male sterility induced by SQ-1

The objective of this study is to elucidate the role of H2O2 in wheat male sterility induced by SQ-1, a new kind of chemical hybridizing agent. We investigated the effects of SQ-1 + DMTU and SQ-1 on the content of H2O2, the transcript levels and activities of SOD, POD, CAT, and APX, and the relative male sterility rate and seed setting rate, and starch-KI staining of pollen grain. The results showed that SQ-1 increased production of H2O2, the activity of APX and the transcript levels of POD and APX, and decreased the activities of SOD, POD, and CAT and the transcript levels of SOD and CAT, compared with control. SQ-1 + DMTU could decrease the production of H2O2 and the transcript levels of POD and APX and increase the activities of SOD, POD, and CAT, compared with SQ-1 alone. However, DMTU could only partly reversed the relative male sterility rate and seed setting rate induced by SQ-1. SQ-1 decreased the number of pollen grains dyed black, compared with control. Treatment with SQ-1 + DMTU increased the number of pollen grains dyed black, compared with SQ-1 alone. Our results indicated that the overproduction of H2O2 was one of reasons for the male sterility induced by SQ-1.

Identification of a novel male sterile wheat mutant dms conferring dwarf status and multi-pistils

Plant height and fertility are two important traits of wheat (Triticum aestivum L.), whose mutants are ideal materials for studies on molecular mechanisms of stem and floral organ development. In this study, we identified a dwarf, multi-pistil and male sterile (dms hereafter) wheat mutant from Zhoumai 18. Simple sequence repeat (SSR) marker assay with 181 primer pairs showed that only one locus of GWM148-2B was divergent between Zhoumai 18 and dms. There were three typical phenotypes in the progeny of dms, tall (T; ca. 0.8 m), semi-dwarf (M; ca. 0.6 m) and dwarf (D; under 0.3 m) plants. Morphological investigation indicated that the internode length of M was shortened by about 20–50 mm each; the internode number of D was 2 less than that of T and Zhoumai 18, and its internode length was shorter also. The pollen vigor and hybridization test demonstrated that dms mutant was male sterility. Segregated phenotypes in progeny of M suggested that the multi-pistils and sterility were controlled by one recessive gene locus which was designated as dms temporarily, and the plant height was controlled by a semi-dominant gene locus Dms. Therefore, progeny individuals of the dms had three genotypes, DmsDms for tall plants, Dmsdms for semi-dwarf plants and dmsdms for dwarf plants. The mutant progenies were individually selected and propagated for more than 6 generations, thus a set of near isogenic lines of T, M and D for dms were developed. This study provides a set germplasms for studies on molecular mechanisms of wheat stem and spike development.

Anther proteomic characterization in temperature sensitive Bainong male sterile wheat

Male sterile line is a useful material for hybridization, but its sterility mechanism, especially proteomic profile, is still not entirely clear. In wheat (Triticum aestivum L.), a male sterile Bainong (BNS) genotype whose sterility could be manipulated by temperature was recently selected. We focused on the proteomic profile change of anthers from the male sterile line (SL) and its conversional line (CL). Two-dimensional gel electrophoresis and MALDI-TOF-MS technologies were utilized for proteomic profiles analysis. Differently abundant protein spots (over 2-fold, P < 0.05) were selected for identification analysis. Compared to CL, 24 up-regulated and 23 down-regulated proteins were identified in SL. Protein metabolism-related proteins, which included a ubiquitin-conjugating enzyme E2 (23 kDa) and an ATP-dependent Clp protease proteolytic subunit, were over-accumulated in SL anthers. Alcohol dehydrogenase ADH1A, fructose-bisphosphate aldolase, chloroplast fructose-bisphosphate aldolase, and NADP-dependent malic enzyme were notably down-regulated in SL anthers. Up-regulated prohibitin protein Wph and a translationallycontrolled tumor protein homolog, and down-regulated histone acetyltransferases HAT1 and HAT2, and DNA directed RNA polymerase subunit α were identified in SL anthers. These dramatically changed proteins may play a crucial role in abnormal anther development and pollen abortion in BNS.