Sterile Anthers Research Articles

Comparative Analysis of the Mitochondrial Genome of Eggplant (Solanum melongena L.) to Identify Cytoplasmic Male Sterility Candidate Genes.

Cytoplasmic male sterility (CMS) is important for commercial hybrid seed production. However, it is still not used in eggplant (Solanum melongena L.), and corresponding regulatory genes and mechanisms of action have not been reported. We report CMS line 327A, which was derived from the hybridization between cultivated and wild eggplants. By looking at different stages of anther development under a microscope, we saw that the 327A anther's tapetum layer vacuolized during meiosis, which caused abortion. To investigate the 327A CMS regulatory genes, the mitochondrial genomes of 327A and its maintainer line 327B were assembled de novo. It was found that 15 unique ORFs (Open Reading Frame) were identified in 327A. RT-PCR and RT-QPCAR tests confirmed that orf312a and orf172a, 327A-specific ORFs with a transmembrane domain, were strongly expressed in sterile anthers of 327A. In addition, orf312a has a chimeric structure with the ribosomal protein subunit rpl16. Therefore, orf312a and orf172a can be considered strong candidate genes for CMS. Concurrently, we analyzed the characteristics of CMS to develop a functional molecular marker, CMS312, targeting a future theoretical basis for eggplant CMS three-line molecular breeding.

Analysis of the cytohistological phenotype of anthers in sunflower with ASI-AG3 in the context of the PET1 mytotype and the allelic genotype for the Rf1 gene

Microsporogenesis and male gametogenesis are processes that, although under tight morphogenetic control, can often be affected by deviations of internal or external stimuli causing androsterility. The article examines the cytohistological phenotypes of sunflower anthers with gibberellin induced androsterility in plants with different mitotype and allelic genotype for the Rf1 gene, in comparison with the cytohistological phenotypes of fertile and sterile anthers (ASC-PET1). The treatment with gibberellic acid (AG3, 0.01%) was carried out by spraying the inflorescence at the budding stage. Cytological samples and those with semi-thin anther sections were analysed by light microscopy. It was determined that the gametocidal effects induced by AG3 associated with degenerative and early apoptotic events in tapetum cells were more severe in plants with sterile cytoplasm and nuclear Rf1 genes compared to those with fertile cytoplasm and no fertility-restoring genes. In plants with ASC, the exogenous gibberellin stimulus does not cause any visible compensatory events of the sterile phenotype, on the contrary, the effects of microsporogenesis inhibition are more temporally advanced, with sporogenous and sporophytic tissue cells showing various lesions in the premeiosis. The general common and differentiation characteristics found at the androsteril phenotypes suggest interactions between orfH522 and Rf1 gene expression products of negative feedback type in untreated plants and positive feedback type in AG3-treated ones. The research is relevant in the context of elucidating the physiological regulatory processes of the sensitivity of male gametophyte development to various stress conditions, which prospectively makes possible regulatory intervention in plants with different androsterility/androfertility control systems.

Morphobiological features of generative organs of fertile and sterile table beet plants and their variability as a result of self-pollination (review)

Beet is considered a difficult breeding object due to its biological peculiarities that make it difficult to obtain hybrid seed progeny. In cross-pollinated crops, the maximum hybridization is achieved by using sterile mother plants, which are extremely rare in the nature. Inbreeding is one of the ways to isolate genotypes with cytoplasmic male sterility, which are controlled by recessive alleles of nuclear genes and S-factor of mitochondrial DNA, from a complex heterogeneous population. The main distinguishing features allowing to identify plants with male sterility are the structure and coloring of anthers, and the quality composition of pollen. The diversity of phenotypic manifestation of marker coloration in sterile anthers is determined by the ratio of different pigments. Diversity of pollen grains of fertile and sterile plants is caused by differences in their development at the late stages of androgenesis. The degree of sterilization varies in different genotypes, which is associated with complex regulatory mechanisms of interaction between the genetic apparatus of the nucleus and the cytoplasm. Along with abnormalities of pollen grain micropopulations, self-fertilization results in morphological changes in the structure of inflorescences and number of flower organs, which has a direct impact on the seed productivity of plants. The studies summarized in this publication are relevant and essential for the search of effective ways to control plant development in ontogenesis and universal selection criteria in the process of creating fundamentally new forms of plants, which is especially important for breeding.

Development differs between independently evolved staminode whorls in the same flower.

Staminodes are commonly studied in hermaphroditic flowers, in which a fraction of the androecium evolves into infertile structures, but few studies have addressed the evolution of staminodes as they occur through the loss of stamen function in carpellate flowers. Plants of Paronychia (Caryophyllaceae) are monoecious with hermaphroditic flowers with one staminodial whorl, except for the dioecious P. chartacea and P. minima. Dioecious species have carpellate flowers that evolved an additional whorl of staminodes, providing an exceptional opportunity to study a second origin of staminodes in the same flower. Using scanning electron microscopy, we observed the development of carpellate and staminate flowers to determine whether the developmental pathway of the staminodes in hermaphroditic flowers was co-opted during the evolutionary transition to unisexual flowers. In carpellate flowers, antesepalous staminodes initiate as sterile anthers that develop similar to functioning stamens, but arrest before full development, leaving a rudimentary anther with lateral lobes that correspond to thecae. After antesepalous staminodes arrest, alternisepalous staminodes initiate as structures that correspond with filaments, as they do in staminate and hermaphroditic flowers. The second origin of staminodes in carpellate flowers evolved using a different developmental pathway than what had previously evolved in the alternisepalous whorl. The two androecial whorls in the same flowers are serialogous as members of the androecium, but are paralogous as staminodes on the basis of structural and developmental differences.

A 163-bp insertion in the Capana10g000198 encoding a MYB transcription factor causes male sterility in pepper (Capsicum annuum L.).

Male sterility provides an efficient approach for commercial exploitation of heterosis. Despite more than 20 genic male sterile (GMS) mutants documented in pepper (Capsicum annuum L.), only two causal genes have been successfully identified. Here, a novel spontaneous recessive GMS mutant, designated msc-3, is identified and characterized at both phenotypic and histological levels. Pollen abortion of msc-3 mutant may be due to the delayed tapetum degradation, leading to the non-degeneration of tetrads callosic wall. Then, a modified MutMap method and molecular marker linkage analysis were employed to fine mapping the msc-3 locus, which was delimited to the ~139.91-kb region harboring 10 annotated genes. Gene expression and structure variation analyses indicate the Capana10g000198, encoding a R2R3-MYB transcription factor, is the best candidate gene for the msc-3 locus. Expression profiling analysis shows the Capana10g000198 is an anther-specific gene, and a 163-bp insertion in the Capana10g000198 is highly correlated with the male sterile (MS) phenotype. Additionally, downregulation of Capana10g000198 in male fertile plants through virus-induced gene silencing resulted in male sterility. Finally, possible regulatory relationships of the msc-3 gene with the other two reported pepper GMS genes, msc-1 and msc-2, have been studied, and comparative transcriptome analysis reveals the expression of 16 GMS homologs are significantly downregulated in the MS anthers. Overall, our results reveal that Capana10g000198 is the causal gene underlying the msc-3 locus, providing important theoretical clues and basis for further in-depth study on the regulatory mechanisms of pollen development in pepper.

Methods of creating substituted lines for hybrid selection of sugar beet: plants of the species Beta vulgaris L. with new sterile cytoplasm of Beta patula L.

Purpose. Due to changing climatic conditions and the spread of new viral diseases, an important issue in beet growing is the expansion of plasma ulture using new sterile cytoplasm from wild species of the genus Beta L.
 as an alternative to Owen’s S-cytoplasm. The goal of this work was to obtain new breeding genotypes based on new cytoplasm. Methods. Cytological, biotechnological. field and laboratory. Results. Availability of recombination in the locus of grafted genes of one-year and two-year development cycles B+b- and hypocotyl colour R+r- was determined using the method of analytical crosses. Earlier we have proved the positive effect of sterility maintainer Beta vulgaris Nxxzz in replacement of the sterility maintainer genome based on our results on the selection of hypocotyl of green color r-rand two-year development cycle b-b- in the interspecific hybrids F2 obtained under the hybridization scheme Beta maritima L: ♀B1C maritima / ♂ Beta vulgaris Nxxzz (Роїк М. В., 2013). In the case of interspecific hybrids, in the conditions of sterile cytoplasm Beta patula L., only backcross offspring of the third cycle of saturating crosses were identified by one- and two-year developmental cycles. Seed bearing plants of BC3S patula of a two-year development cycle were characterized by spreading stems, multi-seeded fruits (3, 4 seeds) and sterile anthers. At the same time, monogermity and a two-year development cycle were restored against the background of sterile cytoplasm of Beta patula L. only among the seed bearing plants of the fourth cycle of saturating crosses. Recombinant genotypes with a red hypocotyl colour and a two-year development cycle BC4S patula R+r- were characterized by multigermity, chimericalness by sterility of CMS‑1 and CMS‑0 type and single- and multi-stem morphological types of anthers. The expression of CMS nuclear genes in the newly replaced BC5S patula lines was marked by high sterility rates of the CMS‑0 type; however, the fertility rates required additional rejection of multigerm seed plants. This is an indicator of the complex structure of the fertility locus in wild Beta patula L. and the special response of the nuclear genome of the sterility maintainers to the new cytoplasmic background. Experimental triploid hybrids with the new introductory sterile cytoplasm Beta patula L. were determined by high sugar content and dry matter content up to 29%. Conclusions. Alloplasmic lines based on the cytoplasm of the wild species Beta patula L are created. Their sugar content at the level of experimental triploid hybrids was 19.5% and 20.5% with a dry matter content of 25% —29%. Alloplasmic lines with a new BCC Beta patula L. recommended for use in the selection process of sugar beets.

Genome-wide identification of cystathionine beta synthase genes in wheat and its relationship with anther male sterility under heat stress.

Cystathionine beta synthase (CBS) domains containing proteins (CDCPs) plays an important role in plant development through regulation of the thioredoxin system, as well as its ability to respond to biotic and abiotic stress conditions. Despite this, no systematic study has examined the wheat CBS gene family and its relation to high temperature-induced male sterility. In this study, 66 CBS family members were identified in the wheat genome, and their gene or protein sequences were used for subsequent analysis. The TaCBS gene family was found to be unevenly distributed on 21 chromosomes, and they were classified into four subgroups according to their gene structure and phylogeny. The results of collinearity analysis showed that there were 25 shared orthologous genes between wheat, rice and Brachypodium distachyon, and one shared orthologous gene between wheat, millet and barley. The cis-regulatory elements of the TaCBS were related to JA, IAA, MYB, etc. GO and KEGG pathway analysis identified these TaCBS genes to be associated with pollination, reproduction, and signaling and cellular processes, respectively. A heatmap of wheat plants based on transcriptome data showed that TaCBS genes were expressed to a higher extent in spikelets relative to other tissues. In addition, 29 putative tae-miRNAs were identified, targeting 41 TaCBS genes. Moreover, qRT-PCR validation of six TaCBS genes indicated their critical role in anther development, as five of them were expressed at lower levels in heat-stressed male sterile anthers than in Normal anthers. Together with anther phenotypes, paraffin sections, starch potassium iodide staining, and qRT-PCR data, we hypothesized that the TaCBS gene has a very important connection with the heat-stressed sterility process in wheat, and these data provide a basis for further insight into their relationship.

FaesAP3_1 Regulates the FaesELF3 Gene Involved in Filament-Length Determination of Long-Homostyle Fagopyrum esculentum.

The identification downstream genes of floral organ identity regulators are critical to revealing the molecular mechanisms underlying floral morphogenesis. However, a general regulatory pathway between floral organ identity genes and their downstream targets is still unclear because of the lack of studies in nonmodel species. Here, we screened a direct downstream target gene, FaesELF3, of a stamen identity transcription factor, FaesAP3_1, in long-homostyle (LH) Fagopyrum esculentum moench by using yeast one-hybrid (Y1H) and dual-luciferase reporter (DR) assays. Furthermore, FaesAP3_1-silenced LH plants that produced flowers with part stamens or anthers homeotically converted into a tepaloid structure, and FaesELF3-silenced plants that had flowers with part stamens consisting of a short filament and empty anther (male sterile anther). All these suggested that transcription factor (TF) FaesAP3_1 directly activates FaesELF3 in order to regulate filament elongation and pollen grain development in LH buckwheat. Our data also suggested that other stamen development pathways independent of FaesAP3_1 remain in F. esculentum.

Combined analysis of transcriptome and metabolome reveals that sugar, lipid, and phenylpropane metabolism are essential for male fertility in temperature-induced male sterile rice.

Photoperiod- and thermosensitive genic male sterility (PTGMS) rice is a vital germplasm resource consisting of two-line hybrid rice in which light and temperature strictly control their fertility changes. Variable environmental conditions present huge risks to the two-lines hybrid seed production. Explaining the regulatory mechanism of male fertility in rice PTGMS lines is an essential prerequisite to ensuring food security production. A group of near-isogenic lines (NILs) of a rice PTGMS line unique to this research group was used for this study. These lines have the same genetic background and regulate male fertility by responding to different temperature changes. Transcriptomic analysis revealed that 315 upregulated genes and 391 regulated genes regulated male fertility in response to temperature changes, and differentially expressed genes (DEGs) were mainly characterized in enrichment analysis as having roles in the metabolic pathways of sugar, lipid and phenylpropanoid. Electron microscopy analysis revealed that a lack of starch accumulation in sterile pollen grains induced by high temperature, with an abnormal exine development and a lack of inner pollen grains. Defective processes for sporopollenin synthesis, sporopollenin transport and pollen wall formation in sterile anthers were verified using qPCR. Targeted metabolomics analysis revealed that most lipids (phospholipids, sphingolipids and fatty acids) and flavonoids (flavones and flavanones) were upregulated in fertile anthers and involved in pollen wall development and male fertility formation, while lignin G units and C-type lignin were the major contributors to pollen wall development. The coding genes for trehalose 6-phosphate phosphatase, beta-1,3-glucanase, phospholipase D and 4-coumarate-CoA ligase are considered essential regulators in the process of male fertility formation. In conclusion, our results indicated that the expression of critical genes and accumulation of metabolites in the metabolism of sugar, lipid, and phenylpropanoid are essential for male fertility formation. The results provide new insights for addressing the negative effects of environmental variation on two-line hybrid rice production.

Overexpression of sesame polyketide synthase A leads to abnormal pollen development in Arabidopsis

BackgroundSesame is a great reservoir of bioactive constituents and unique antioxidant components. It is widely used for its nutritional and medicinal value. The expanding demand for sesame seeds is putting pressure on sesame breeders to develop high-yielding varieties. A hybrid breeding strategy based on male sterility is one of the most effective ways to increase the crop yield. To date, little is known about the genes and mechanism underlying sesame male fertility. Therefore, studies are being conducted to identify and functionally characterize key candidate genes involved in sesame pollen development. Polyketide synthases (PKSs) are critical enzymes involved in the biosynthesis of sporopollenin, the primary component of pollen exine. Their in planta functions are being investigated for applications in crop breeding.ResultsIn this study, we cloned the sesame POLYKETIDE SYNTHASE A (SiPKSA) and examined its function in male sterility. SiPKSA was specifically expressed in sesame flower buds, and its expression was significantly higher in sterile sesame anthers than in fertile anthers during the tetrad and microspore development stages. Furthermore, overexpression of SiPKSA in Arabidopsis caused male sterility in transgenic plants. Ultrastructural observation showed that the pollen grains of SiPKSA-overexpressing plants contained few cytoplasmic inclusions and exhibited an abnormal pollen wall structure, with a thicker exine layer compared to the wild type. In agreement with this, the expression of a set of sporopollenin biosynthesis-related genes and the contents of their fatty acids and phenolics were significantly altered in anthers of SiPKSA-overexpressing plants compared with wild type during anther development.ConclusionThese findings highlighted that overexpression of SiPKSA in Arabidopsis might cause male sterility through defective pollen wall formation. Moreover, they suggested that SiPKSA modulates vibrant pollen development via sporopollenin biosynthesis, and a defect in its regulation may induce male sterility. Therefore, genetic manipulation of SiPKSA might promote hybrid breeding in sesame and other crop species.

Comparative Transcriptome Analysis of Male Sterile Anthers Induced by High Temperature in Wheat (Triticum aestivum L.).

Global warming will have a negative effect on agricultural production as high temperature (HT) stress can seriously threaten plant growth and reproduction. Male sterility caused by HT may be exploited by the creation of a male-sterile line, which has great potential for application in crop heterosis. Therefore, it is important to understand the molecular mechanisms of anther abortion induced by HT in wheat, which remain unclear at present. In this study, we performed phenotype improve language in the abstract and comparative transcriptome analysis of the male sterile anthers induced by HT in wheat. Compared with Normal anthers, the cytological analysis indicated that HT-induced male sterile anthers were smaller and had no starch accumulation in pollen grains, which is consistent with the results observed by scanning electron microscopy (SEM). The 9601 differentially expressed genes (DEGs) identified by transcriptome sequencing compared with the Normal anthers were noticeably involved in the following pathways: starch and sucrose metabolism, phosphatidylinositol (PI) signaling system, peroxidase activity and response to oxidative stress, and heme binding. In addition, TUNEL assays were performed and the results further confirmed the excessive accumulation of reactive oxygen species (ROS) in sterile anthers. Moreover, a total of 38 hub genes were obtained from the protein-protein interaction network analysis of these pathways, including genes, for example, heat shock protein 90 (HSP90), thioredoxin-like protein 1, peroxidase (POD), calreticulin, UDP glucose pyrophosphorylase (UGPase), sucrose synthase, phosphatidylinositol-4-phosphate 5-Kinase (PIP5K), cytochrome c, and Cystathionine beta-synthase X6-like (CBSX6-like). These findings provide insights for predicting the functions of the candidate genes, and the comprehensive analysis of our results is helpful for studying the abortive interaction mechanism induced by HT in wheat.

Metabolomic profiling of SQ-1-induced changes in starch metabolism in sterile anthers of wheat

Metabolomic profiling of SQ-1-induced changes in starch metabolism in sterile anthers of wheat

Unraveling the Genetic Basis of Fertility Restoration for Cytoplasmic Male Sterile Line WNJ01A Originated From Brassica juncea in Brassica napus.

Crosses that lead to heterosis have been widely used in the rapeseed (Brassica napus L.) industry. Cytoplasmic male sterility (CMS)/restorer-of-fertility (Rf) systems represent one of the most useful tools for rapeseed production. Several CMS types and their restorer lines have been identified in rapeseed, but there are few studies on the mechanisms underlying fertility restoration. Here, we performed morphological observation, map-based cloning, and transcriptomic analysis of the F2 population developed by crossing the CMS line WNJ01A with its restorer line Hui01. Paraffin-embedded sections showed that the sporogenous cell stage was the critical pollen degeneration period, with major sporogenous cells displaying loose and irregular arrangement in sterile anthers. Most mitochondrial electron transport chain (mtETC) complex genes were upregulated in fertile compared to sterile buds. Using bulked segregant analysis (BSA)-seq to analyze mixed DNA pools from sterile and fertile F2 buds, respectively, we identified a 6.25 Mb candidate interval where Rfw is located. Using map-based cloning experiments combined with bacterial artificial chromosome (BAC) clone sequencing, the candidate interval was reduced to 99.75 kb and two pentatricopeptide repeat (PPR) genes were found among 28 predicted genes in this interval. Transcriptome sequencing showed that there were 1679 DEGs (1023 upregulated and 656 downregulated) in fertile compared to sterile F2 buds. The upregulated differentially expressed genes (DEGs) were enriched in the Kyoto Encyclopedia of Genes and Genomes (KEGG) lysine degradation pathway and phenylalanine metabolism, and the downregulated DEGs were enriched in cutin, suberine, and wax biosynthesis. Furthermore, 44 DEGs were involved in pollen and anther development, such as tapetum, microspores, and pollen wall development. All of them were upregulated except a few such as POE1 genes (which encode Pollen Ole e I allergen and extensin family proteins). There were 261 specifically expressed DEGs (9 and 252 in sterile and fertile buds, respectively). Regarding the fertile bud-specific upregulated DEGs, the ubiquitin–proteasome pathway was enriched. The top four hub genes in the protein–protein interaction network (BnaA09g56400D, BnaA10g18210D, BnaA10g18220D, and BnaC09g41740D) encode RAD23d proteins, which deliver ubiquitinated substrates to the 26S proteasome. These findings provide evidence on the pathways regulated by Rfw and improve our understanding of fertility restoration.

GmMs1 encodes a kinesin-like protein essential for male fertility in soybean (Glycine max L.).

The application of heterosis is a promising approach for greatly increasing yield in soybean (Glycine max L.). Nuclear male sterility is essential for hybrid seed production and the utilization of heterosis. Here we report the cloning of the gene underlying the soybean male-sterile mutant ms-1, which has been widely used for recurrent selection in soybean breeding programs. We initially delimited the ms1 locus to a 16.15 kb region on chromosome 13, based on SLAF_BSA sequencing followed by genotyping of an F2 population segregating for the locus. Compared with the same region in fertile plants, the mutant region lacks a sequence of approximately 38.7 kb containing five protein-coding genes, including an ortholog of the kinesin-like protein gene NACK2, named GmMs1. The GmMs1 knockout plants generated via CRISPR/Cas-mediated gene editing displayed a complete male-sterile phenotype. Metabolic profiling showed that fertile anthers accumulated starch and sucrose normally, whereas sterile anthers had higher anthocyanin levels and lower flavonoid levels and lower antioxidant enzyme activities. These results provide insights into the molecular mechanisms governing male sterility and demonstrate that GmMs1 could be used to create male-sterile lines through targeted mutagenesis. These findings pave the way for designing seed production technology and an intelligent male-sterile line system to utilize heterosis in soybean.

Potential Association of Reactive Oxygen Species With Male Sterility in Peach.

Male sterility is an important agronomic trait for hybrid vigor utilization and hybrid seed production, but its underlying mechanisms remain to be uncovered. Here, we investigated the mechanisms of male sterility in peach using a combined cytology, physiology, and molecular approach. Cytological features of male sterility include deformed microspores and tapetum cells along with absence of pollen grains. Microspores had smaller nucleus at the mononuclear stage and were compressed into belts and subsequently disappeared in the anther cavity, whereas tapetum cells were swollen and vacuolated, with a delayed degradation to flowering time. Male sterile anthers had an ROS burst and lower levels of major antioxidants, which may cause abnormal development of microspores and tapetum, leading to male sterility in peach. In addition, the male sterility appears to be cytoplasmic in peach, which could be due to sequence variation in the mitochondrial genome. Our results are helpful for further investigation of the genetic mechanisms underlying male sterility in peach.

Dynamic expression of miRNAs and functional analysis of target genes involved in the response to male sterility of the wheat line YS3038

Thermosensitive cytoplasmic male sterile (TCMS) lines play an important role in wheat breeding, heterosis utilization, and germplasm innovation. MicroRNAs (miRNAs) can regulate the expression level of target genes by inhibiting the translation of these genes. YS3038 is a wheat TCMS line. In this study, the fertility conversion mechanism of YS3038 was studied by examining the abortion characteristics of YS3038, the regulation pattern of miRNAs and the target genes of miRNAs in YS3038. MiRNA-seq was performed on three important stages of YS3038 under sterile and fertile conditions. Then, the clean reads were aligned with some databases to filter other ncRNAs and repeats. The known miRNAs and novel miRNAs were predicted by sequence comparison with known miRNAs from miRbase. Differential expression of miRNAs between different stages and between different fertile conditions was analyzed, and functional analysis of target genes with opposite expression patterns as those of the miRNAs was conducted. The Ubisch bodies and microspores of sterile anthers were covered with filamentous materials. The degradation of the tapetum cells, the chloroplast structure of endothecium cells, and the microspore structure were abnormal. Microspore development was hindered from the late uninucleate stage to the binucleate stage. Twenty, 52, and 68 differentially expressed miRNAs (DEmiRs) were identified at the early uninucleate, late uninucleate, and binucleate stages, respectively, and there were 0, 7, and 72 differentially expressed target genes (DETGs), respectively, at these three stages. At the binucleate stage, 29 DEmiRs had 41 target mRNAs in total, and the expression patterns of the 41 target mRNAs were opposite to those of the 29 miRNAs. Fifteen significantly enriched KEGG pathways were associated with the 41 target mRNAs. Leucine-rich repeat receptor-like kinases (LRR-RLKs) play important roles in plant developmental and physiological processes. Some studies have shown that the expression of LRR-RLKs is related to the differentiation of microsporocytes and tapetum cells and to male sterility. An LRR-RLK (TaeRPK) gene was silenced by the barley stripe mosaic virus-induced gene silencing (BSMV-VIGS) method, and the seed setting rates of the TaeRPK-silenced plants (3.51%) were significantly lower than those of the negative control plants (88.78%) (P < 0.01). Thus, the TaeRPK gene is likely to be involved in the fertility conversion of YS3038.

TMT-based quantitative proteomics analyses of sterile/fertile anthers from a genic male-sterile line and its maintainer in cotton (Gossypium hirsutum L.)

Genetic male sterility (GMS) in cotton is important for utilization of heterosis. However, the molecular mechanism of GMS is poorly known. In this study, cytological and proteomics analyses of anthers were conducted in three stages (stage 3 to 5) between GMS line (GA18) and its maintainer (GA18M). The cross-section observation revealed that the tapetal layer in stage 3 was thinner in GA18 compared to GA18M, and the tapetum cells did not degrade in stage 4 in GA18, thus providing no material for microspore development. A total of 1952 differentially expressed proteins (DEPs) were identified between GA18 and GA18M anthers. They were annotated to 52 gene ontology (GO) terms and enriched in 115 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, which formed several complex regulator networks, and dozens of important nodes were identified. Moreover, DEPs were also identified between two consecutive stages of GA18 and GA18M, with functional analyses indicating that numerous developmental differences existed between fertile and sterile anthers. The metabolic pathways were significantly altered, including decreased carbohydrate metabolism, ribosome defects, disturbed protein synthesis, disrupted flavonoids synthesis, etc., that may be involved in male sterility. Overall, these results provide genetic resources that help decipher the molecular mechanisms behind GMS. SignificanceMale sterility is a common phenomenon in flowering plant species, and plays a role in the application of heterosis. However, the molecular mechanism of it remains to be elucidated. Using cytological and proteomics approaches, we found that the tapetal layer development retardation may be the reason of male sterility, which was different from the delayed degradation described in previous studies. More than one thousand differentially expressed proteins were identified between male sterile line and its maintainer, forming a complex regulatory network, and the key nodes were remarked that could be used as candidate proteins related to male sterility in future study. Dozens of metabolic pathways were significantly altered, among them, ribosome defects was a novel pathway that may be involved in male sterility. These results enhance our understanding of the molecular mechanism governing male sterility and lay a foundation for clone of genes association with male sterility.

Cytological and genetic characterisation of dominant GMS line Shaan-GMS in Brassica napus L.

Genic male sterility (GMS) is an effective pollination control system applied in the hybrid breeding of Brassica napus L. Shaan-GMS is a spontaneous mutant of dominant GMS in B. napus. In this research, anther abortion in the homozygous two-type line 9A15AB derived from Shaan-GMS was characterised with the combined use of light microscopy and transmission electron microscopy. Results indicated that the most striking differences between the fertile and sterile plants occurred in the tapetum in the early microsporocyte stage. In sterile plants, the tapetal cells were irregularly arranged, multi-layered and occupied the growing space of microsporocytes. When entering into meiosis, the tapetum cells degraded and the cytoplasm fused. Some oval monolayer or bilayer membrane organelles existed in the tapetal cells in sterile anthers. Mitochondria in the tapetal cells were abnormal, and middle layer cells degraded early. Pollen mother cells of Shaan-GMS degenerated at the start of meiosis and ceased at the anaphase I stage, with no dyads or tetrads formed. The combined effects of the abnormal development of the tapetum, the middle layer cells and meiosis lead to male sterility in Shaan-GMS. Inheritance of male sterility of Shaan-GMS is controlled by a monogenically multiallelic locus with three different alleles (Ms, ms and Mf), with a relationship expressed as Mf > Ms and Ms > ms. The findings help lay the foundation for illustrating the mechanism of male sterility and the utilisation of Shaan-GMS in rapeseed.

Multiomics approach unravels fertility transition in a pigeonpea line for a two-line hybrid system.

Pigeonpea [Cajanus cajan (L.) Millsp.] is a pulse crop cultivated in the semi-arid regions of Asia and Africa. It is a rich source of protein and capable of alleviating malnutrition, improving soil health and the livelihoods of small-holder farmers. Hybrid breeding has provided remarkable improvements for pigeonpea productivity, but owing to a tedious and costly seed production system, an alternative two-line hybrid technology is being explored. In this regard, an environment-sensitive male sterile line has been characterized as a thermosensitive male sterile line in pigeonpea precisely responding to day temperature. The male sterile and fertile anthers from five developmental stages were studied by integrating transcriptomics, proteomics and metabolomics supported by precise phenotyping and scanning electron microscopic study. Spatio-temporal analysis of anther transcriptome and proteome revealed 17 repressed DEGs/DEPs in sterile anthers that play a critical role in normal cell wall morphogenesis and tapetal cell development. The male fertility to sterility transition was mainly due to a perturbation in auxin homeostasis, leading to impaired cell wall modification and sugar transport. Limited nutrient utilization thus leads to microspore starvation in response to moderately elevated day temperature which could be restored with auxin-treatment in the male sterile line. Our findings outline a molecular mechanism that underpins fertility transition responses thereby providing a process-oriented two-line hybrid breeding framework for pigeonpea.

Comparative transcriptome analysis indicates that a core transcriptional network mediates isonuclear alloplasmic male sterility in wheat (Triticum aestivum L.)

BackgroundCytoplasmic male sterility (CMS) plays a crucial role in the utilization of heterosis and various types of CMS often have different abortion mechanisms. Therefore, it is important to understand the molecular mechanisms related to anther abortion in wheat, which remain unclear at present.ResultsIn this study, five isonuclear alloplasmic male sterile lines (IAMSLs) and their maintainer were investigated. Cytological analysis indicated that the abortion type was identical in IAMSLs, typical and stainable abortion, and the key abortive period was in the binucleate stage. Most of the 1,281 core shared differentially expressed genes identified by transcriptome sequencing compared with the maintainer in the vital abortive stage were involved in the metabolism of sugars, oxidative phosphorylation, phenylpropane biosynthesis, and phosphatidylinositol signaling, and they were downregulated in the IAMSLs. Key candidate genes encoding chalcone--flavonone isomerase, pectinesterase, and UDP-glucose pyrophosphorylase were screened and identified. Moreover, further verification elucidated that due to the impact of downregulated genes in these pathways, the male sterile anthers were deficient in sugar and energy, with excessive accumulations of ROS, blocked sporopollenin synthesis, and abnormal tapetum degradation.ConclusionsThrough comparative transcriptome analysis, an intriguing core transcriptome-mediated male-sterility network was proposed and constructed for wheat and inferred that the downregulation of genes in important pathways may ultimately stunt the formation of the pollen outer wall in IAMSLs. These findings provide insights for predicting the functions of the candidate genes, and the comprehensive analysis of our results was helpful for studying the abortive interaction mechanism in CMS wheat.