Male Sterile Mutant Research Articles

Floral Developmental Morphology and Biochemical Characteristics of Male Sterile Mutants of Lagerstroemia indica

Male sterility is a common phenomenon in higher plants and often plays an important role in the selection of superior offspring. ‘Xiang Yun’ is a mutant of Lagerstroemia indica that does not bear fruit after flowering, and its flowering period is significantly longer than that of normal L. indica. To explore the timing and molecular mechanisms of sterility in ‘Xiang Yun’, this study determined the period of sterility through anatomical observation and compared the content of nutrients and the activity of antioxidative enzymes at different stages of flower development. Finally, sequence alignment and qPCR were used to analyze the differences in pollen development genes between ‘Xiang Yun’ and ‘Hong Ye’. The results showed that the anthers of ‘Xiang Yun’ dispersed pollen normally, but the pollen grains could not germinate normally. Observations with scanning electron microscopy revealed that the pollen grains were uneven in size and shriveled in shape. Further observation of anther sections found that abnormal development of the microspores began at the S2 stage, with the callose wall between microspores of ‘Xiang Yun’ being thicker than that of ‘Hong Ye’. In addition, during the flower development of ‘Xiang Yun’, the contents of soluble sugar, soluble protein, free proline, and triglycerides were deficient to varying degrees, and the activities of POD, SOD, and MDA were lower. Sequence alignment and qPCR showed that there were several mutations in EFD1, TPD1, and DEX1 of ‘Xiang Yun’ compared with ‘Hong Ye’, and the expression levels of these genes were abnormally elevated in the later stages of development. Our results clarified the timing and phenotype of male sterility in ‘Xiang Yun’. This provides solid and valuable information for further research on the molecular mechanism of sterility in ‘Xiang Yun’ and the genetic breeding of crape myrtle.

An explanation for the prevalence of XY over ZW sex determination in species derived from hermaphroditism

The many independent transitions from hermaphroditism to separate sexes (dioecy) in flowering plants and some animal clades must often have involved the emergence of a heterogametic sex-determining locus, the basis of XY and ZW sex determination (i.e., male and female heterogamety). Current estimates indicate that XY sex determination is much more frequent than ZW, but the reasons for this asymmetry are unclear. One proposition is that separate sexes evolve through the invasion of sterility mutations at closely linked loci, in which case XY sex determination evolves if the initial male sterility mutation is fully recessive. Alternatively, dioecy may evolve via the gradual divergence of male and female phenotypes, but the genetic basis of such divergence and its connection to XY and ZW systems remain poorly understood. Using mathematical modeling, we show how dioecy with XY or ZW sex determination can emerge from the joint evolution of resource allocation to male and female function with its genetic architecture. Our model reveals that whether XY or ZW sex determination evolves depends on the trade-off between allocation to male and female function, and on the mating system of the ancestral hermaphrodites, with selection for female specialization or inbreeding avoidance both favoring XY sex determination. Together, our results cast light on an important but poorly understood path from hermaphroditism to dioecy, and provide an adaptive hypothesis for the preponderance of XY systems. Beyond sex and sex determination, our model shows how ecology can influence the way selection shapes the genetic architecture of polymorphic traits.

GLUTAMYL-tRNA SYNTHETASE 1 deficiency confers thermosensitive male sterility in rice by affecting reactive oxygen species homeostasis.

Temperature is one of the key environmental factors influencing crop fertility and yield. Understanding how plants sense and respond to temperature changes is, therefore, crucial for improving agricultural production. In this study, we characterized a temperature-sensitive male sterile mutant in rice (Oryza sativa), glutamyl-tRNA synthetase 1-2 (ers1-2), that shows reduced fertility at high temperatures and restored fertility at low temperatures. Mutation of ERS1 resulted in severely delayed pollen development and meiotic progression at high temperatures, eventually leading to male sterility. Moreover, meiosis-specific events, including synapsis and crossover formation, were also delayed in ers1-2 compared with the wild type. However, these defects were all mitigated by growing ers1-2 at low temperatures. Transcriptome analysis and measurement of ascorbate, glutathione, and hydrogen peroxide (H2O2) contents revealed that the delayed meiotic progression and male sterility in ers1-2 were strongly associated with changes in reactive oxygen species (ROS) homeostasis. At high temperatures, ers1-2 exhibited decreased accumulation of ROS scavengers and overaccumulation of ROS. In contrast, at low temperatures, the antioxidant system of ROS was more active, and ROS contents were lower. These data suggest that ROS homeostasis in ers1-2 is disrupted at high temperatures but restored at low temperatures. We speculate that ERS1 dysfunction leads to changes in ROS homeostasis under different conditions, resulting in delayed or rescued meiotic progression and thermosensitive male fertility. ers1-2 may hold great potential as a thermosensitive material for crop heterosis breeding.

Isolation and evaluation of novel male sterile and self‐incompatible mutant lines of mungbean (Vigna radiata L.)

AbstractHeterosis has long been harnessed in crop production, and while crop fertility lays the groundwork for leveraging heterosis, there remains a dearth of comprehensive data regarding genic male sterility in mungbean. Mungbean (Vigna radiata (L.) Wilczek), being a self‐pollinating crop, exhibits significant hybrid vigour in F1 hybrid seed yield, suggesting the potential for developing hybrid varieties to propel mungbean yield beyond existing plateaus. A new male sterile mutant was induced, isolated, stabilized and evaluated, via gamma irradiation at a rate of 600 Gy, identified in the M3 generation of the GM 4 accession. A uniform male sterile line was crossbred with the wild‐type parental plant to explore the inheritance pattern of male sterility. Pollen sterility was validated through the acetocarmine test, while stigma viability was ascertained using the hydrogen peroxide. While all F1 plants displayed fertile flowers, the F2 generation showed a clear 3:1 segregation ratio for fertile to male sterile plants, indicating the control of male sterility by a single recessive gene, mms. Also, a novel existence of a truncated stigma nestled within the anther column inhibits the effective reception of pollen during anthesis, offering potential in reducing emasculation time by a slight cut on the flower bud and enhancing cross pollination during hybrid seed production. The isolation of these two mutants is poised to significantly advance the global mungbean hybrid breeding programme.

Cloning and functional verification of the male sterile gene BrQRT3 in Chinese cabbage

Chinese cabbage is a cross-pollinated crop with significant heterosis, and male sterile lines are an important way to produce hybrid seeds. In this study, a male sterile mutant msm0795 was identified in an EMS-mutagenized population of Chinese cabbage. Cytological observations revealed that the microspores failed to separate after the tetrad stage, and thus developed into abnormal pollen grains, resulting in anther abortion. MutMap combined with Kompetitive Allele Specific PCR genotyping showed that BraA01g011280.3.5 C was identified as the candidate gene, which encodes polygalacturonase QRT3 and plays a direct role in the degradation of pollen mother cell wall during microspore development, named BrQRT3. Subcellular localization and expression analyses demonstrated that BrQRT3 was localized in the cell membrane and was ubiquitously expressed in roots, stems, leaves, flower buds, and flowers, but the expression of BrQRT3 was gradually suppressed with the anther development. Ectopic expression confirmed that over-expression of BrQRT3 in qrt3 background Arabidopsis mutant can rescue the pollen defects caused by loss of AtQRT3 function. It is the first time to achieve a male sterile mutant caused by the mutation of BrQRT3 in Chinese cabbage. These findings contribute to elucidate the mechanism of BrQRT3 in regulating stamen development of Chinese cabbage.

Transcriptome analysis reveals the potential lncRNA-mRNA modules involved in genetic male sterility and fertility of Chinese cabbage (brassica rapa L. ssp. pekinensis).

Long non-coding RNAs (lncRNAs) play a crucial role in regulating gene expression vital for the growth and development of plants. Despite this, the role of lncRNAs in Chinese cabbage (Brassica rapa L. ssp. pekinensis) pollen development and male fertility remains poorly understood. In this study, we characterized a recessive genic male sterile mutant (366-2S), where the delayed degradation of tapetum and the failure of tetrad separation primarily led to the inability to form single microspores, resulting in male sterility. To analyze the role of lncRNAs in pollen development, we conducted a comparative lncRNA sequencing using anthers from the male sterile mutant line (366-2S) and the wild-type male fertile line (366-2F). We identified 385 differentially expressed lncRNAs between the 366-2F and 366-2S lines, with 172 of them potentially associated with target genes. To further understand the alterations in mRNA expression and explore potential lncRNA-target genes (mRNAs), we performed comparative mRNA transcriptome analysis in the anthers of 366-2S and 366-2F at two stages. We identified 1,176 differentially expressed mRNAs. Remarkably, GO analysis revealed significant enrichment in five GO terms, most notably involving mRNAs annotated as pectinesterase and polygalacturonase, which play roles in cell wall degradation. The considerable downregulation of these genes might contribute to the delayed degradation of tapetum in 366-2S. Furthermore, we identified 15 lncRNA-mRNA modules through Venn diagram analysis. Among them, MSTRG.9997-BraA04g004630.3C (β-1,3-glucanase) is associated with callose degradation and tetrad separation. Additionally, MSTRG.5212-BraA02g040020.3C (pectinesterase) and MSTRG.13,532-BraA05g030320.3C (pectinesterase) are associated with cell wall degradation of the tapetum, indicating that these three candidate lncRNA-mRNA modules potentially regulate pollen development. This study lays the foundation for understanding the roles of lncRNAs in pollen development and for elucidating their molecular mechanisms in regulating male sterility in Chinese cabbage.

Candidate Gene Identification and Transcriptome Analysis of Tomato male sterile-30 and Functional Marker Development for ms-30 and Its Alleles, ms-33, 7B-1, and stamenless-2.

Male sterility is a valuable trait for hybrid seed production in tomato (Solanum lycopersicum). The mutants male sterile-30 (ms-30) and ms-33 of tomato exhibit twisted stamens, exposed stigmas, and complete male sterility, thus holding potential for application in hybrid seed production. In this study, the ms-30 and ms-33 loci were fine-mapped to 53.3 kb and 111.2 kb intervals, respectively. Tomato PISTILLATA (TPI, syn. SlGLO2), a B-class MADS-box transcription factor gene, was identified as the most likely candidate gene for both loci. TPI is also the candidate gene of tomato male sterile mutant 7B-1 and sl-2. Allelism tests revealed that ms-30, ms-33, 7B-1, and sl-2 were allelic. Sequencing analysis showed sequence alterations in the TPI gene in all these mutants, with ms-30 exhibiting a transversion (G to T) that resulted in a missense mutation (S to I); ms-33 showing a transition (A to T) that led to alternative splicing, resulting in a loss of 46 amino acids in protein; and 7B-1 and sl-2 mutants showing the insertion of an approximately 4.8 kb retrotransposon. On the basis of these sequence alterations, a Kompetitive Allele Specific PCR marker, a sequencing marker, and an Insertion/Deletion marker were developed. Phenotypic analysis of the TPI gene-edited mutants and allelism tests indicated that the gene TPI is responsible for ms-30 and its alleles. Transcriptome analysis of ms-30 and quantitative RT-PCR revealed some differentially expressed genes associated with stamen and carpel development. These findings will aid in the marker-assisted selection for ms-30 and its alleles in tomato breeding and support the functional analysis of the TPI gene.

Exploitation of Male Sterility for Heterosis Breeding in Vegetable Crops

The phenomenon of heterosis is extensively utilized in vegetable crops for the exploitation of hybrid vigor. Presently, hybrids are popular in majority of the vegetable crops due to their uniform shape, size, color and quality, providing better returns to the growers. The main worry regarding production of hybrid seeds is the scarcity and high cost of specialized labor, which increasingly raises the cost of hybrid seed production, especially for solanaceous crops, where manual emasculation and pollination mechanisms still play an essential role. Thus, there is need to exploit different mechanisms which results in decreasing the cost of hybrids. Among these mechanisms, male sterility is very efficient and economic method for the large-scale F1 seed production. Many male sterile mutants have also been discovered which eliminates the need of tedious emasculation process. Undesirable type of plants at the early stage of growth can be identified by using certain marker genes and thus facilitates the production of hybrid seeds. Exploiting heterosis in vegetable crops is more profitable as it increases the uniformity, resistance against various insect-pests and yield.

Mutations in BrABCG26, encoding an ATP-binding cassette transporter, are responsible for male sterility in Chinese cabbage (Brassica rapa L. ssp. pekinensis).

The gene BrABCG26 responsible for male sterility of Chinese cabbage was confirmed by two allelic mutants. Male-sterile lines are an important way of heterosis utilization in Chinese cabbage. In this study, two allelic male-sterile mutants msm3-1 and msm3-2 were obtained from a Chinese cabbage double haploid (DH) line 'FT' by using EMS-mutagenesis. Compared to the wild-type 'FT,' the stamens of mutants were completely degenerated and had no pollen, and other characters had no obvious differences. Cytological observation revealed that the failure of vacuolation of the mononuclear microspore, accompanied by abnormal tapetal degradation, resulted in anther abortion in mutants. Genetic analysis showed thata recessive gene controlled the mutant trait. MutMap combined with kompetitive allele specific PCR genotyping analyses showed that BraA01g038270.3C, encoding a transporter ABCG26 that played a vital role in pollen wall formation, was the candidate gene for msm3-1, named BrABCG26. Compared with wild-type 'FT,' the mutations existed on the second exon (C to T) and the sixth exon (C to T) of BrABCG26 gene in mutants msm3-1 and msm3-2, leading to the loss-of-function truncated protein, which verified the BrABCG26 function in stamen development. Subcellular localization and expression pattern analysis indicated that BrABCG26 was localized in the nucleus and was expressed in all organs, with the highest expression in flower buds. Compared to the wild-type 'FT,' the expressions of BrABCG26 were significantly reduced in flower buds and anthers of mutants. Promoter activity analysis showed that a strong GUS signal was detected in flower buds. These results indicated that BrABCG26 is responsible for the male sterility of msm3 mutants in Chinese cabbage.

Characterization of the CMS genetic regulation through comparative complete mitochondrial genome sequencing in Nicotiana tabacum.

Mitochondrial genomes (mitogenomes) of flowering plants vary greatly in structure and size, which can lead to frequent gene mutation, rearrangement, or recombination, then result in the cytoplasmic male sterile (CMS) mutants. In tobacco (Nicotiana tabacum), suaCMS lines are widely used in heterosis breeding; however, the related genetic regulations are not very clear. In this study, the cytological observation indicated that the pollen abortion of tobacco suaCMS(HD) occurred at the very early stage of the stamen primordia differentiation. In this study, the complete mitochondrial genomes of suaCMS(HD) and its maintainer HD were sequenced using the PacBio and Illumina Hiseq technology. The total length of the assembled mitogenomes of suaCMS(HD) and HD was 494,317bp and 430,694bp, respectively. Comparative analysis indicated that the expanded 64 K bases in suaCMS(HD) were mainly located in noncoding regions, and 23 and 21 big syntenic blocks (>5000bp) were found in suaCMS(HD) and HD with a series of repeats. Electron transport chain-related genes were highly conserved in two mitogenomes, except five genes (ATP4, ATP6, COX2, CcmFC, and SDH3) with substantial substitutions. Three suaCMS(HD)-specific genes, orf261, orf291, and orf433, were screened. Sequence analysis and RT-PCR verification showed that they were unique to suaCMS(HD). Further gene location analysis and protein property prediction indicated that all the three genes were likely candidates for suaCMS(HD). This study provides new insight into understanding the suaCMS mechanism and is useful for improving tobacco breeding.

The Assessment of Natural Cross Pollination Properties of a Novel Male-Sterile-Female-Fertile Mutation msLC01 in Soybean.

The value of a novel soybean male-sterile mutation msLC01 in breeding practice was determined by its outcrossing properties. Then, the effects of different planting arrangements on the pod set characteristics of male-sterile plants were assessed by using orthogonal experiments at two sites. At the same time, the effects of msLC01 male sterility on other traits were assessed in two C2F2 populations. In addition, the nectar secretion and natural outcross of male-sterile plants from four msLC01 lines were compared with one ms1 line and one ms6 line. The results of the orthogonal experiment showed that the pod numbers and pod set rates of male-sterile plants were decisively different between the two experimental sites but not between the two levels of the other factors. Both increasing the ratio of paternal parent to maternal parent and planting the parental seeds in a mixed way, the proportion of seeds pollinated by the target parent pollen could be increased. Except for the pod number per plant trait, there was no significant difference between male-sterile plants and their fertile siblings. The amount of nectar significantly differed among the lines. Compared with ms1 and ms6 male-sterile plants, the four msLC01 lines possessed significantly more or similar numbers of pod sets. The results of this study lay a foundation for the future use of this mutant in soybean breeding.

Changes in m6A RNA methylation are associated with male sterility in wolfberry

BackgroundN6-methyladenosine (m6A) modification is the most abundant type of RNA modification in eukaryotic cells, playing pivotal roles in multiple plant growth and development processes. Yet the potential role of m6A in conferring the trait of male sterility in plants remains unknown.ResultsIn this study, we performed RNA-sequencing (RNA-Seq) and m6A-sequencing (m6A-Seq) of RNAs obtained from the anther tissue of two wolfberry lines: ‘Ningqi No.1’ (LB1) and its natural male sterile mutant ‘Ningqi No.5’ (LB5). Based on the newly assembled transcriptome, we established transcriptome-wide m6A maps for LB1 and LB5 at the single nucleus pollen stage. We found that the gene XLOC_021201, a homolog of m6A eraser-related gene ALKBH10 in Arabidopsis thaliana, was significantly differentially expressed between LB1 and LB5. We also identified 1642 and 563 m6A-modified genes with hypermethylated and hypomethylated patterns, respectively, in LB1 compared with LB5. We found the hypermethylated genes significantly enriched in biological processes related to energy metabolism and lipid metabolism, while hypomethylation genes were mainly linked to cell cycle process, gametophyte development, and reproductive process. Among these 2205 differentially m6A methylated genes, 13.74% (303 of 2205) were differentially expressed in LB1 vis-à-vis LB5.ConclusionsThis study constructs the first m6A transcriptome map of wolfberry and establishes an association between m6A and the trait of male sterility in wolfberry.

Identification and functional characterization of Si4CLL1 controlling the male sterility in the sesame genic male sterile mutant, ms1812

AbstractSesame (Sesamum indicum L.), 2n = 2x = 26, is an important oilseed crop with high heterosis potential. In this study, we explored the histological and genetic characters and causal gene of a male sterile sesame mutant, ms1812 (ms3). The microspore abortion of ms1812 that led to male sterility (MS) occurred at the tetrad stage of microspore development. No normal pollens were formed and observed in anther chambers of ms1812 at the microspore mature stage. With the aid of high‐efficient association mapping and the sesame var. Yuzhi 11 reference genome, we identified the single nucleotide polymorphism 10505640 on SiChr.3 segregating with the MS and the mutated gene, Si4cll1, controlling the recessive MS in sesame. Si4CLL1 encodes a 4‐coumarate‐CoA ligase‐like protein. The 1292th nucleotide (T1292) of Si4CLL1 mutated into A1292, resulting in the amino acid change from Ile239 to Lys239. Si4CLL1 is mainly expressed in young developing buds. Compared with the wild type, the transcript abundance of Si4cll1 in the developing buds of ms1812 reduced significantly, which might affect the phenylpropanoid biosynthesis pathway and finally, the lignin formation of microspore cell wall. These findings provide key new information for deciphering the molecular mechanism underlying MS and hybrid variety breeding assisted by MS in sesame.

Evolution of sex-determination in dioecious plants: From active Y to X/A balance?

Sex chromosomes in plants have been known for a century, but only recently have we begun to understand the mechanisms behind sex determination in dioecious plants. Here, we discuss evolution of sex determination, focusing on Silene latifolia, where evolution of separate sexes is consistent with the classic "two mutations" model-a loss of function male sterility mutation and a gain of function gynoecium suppression mutation, which turned an ancestral hermaphroditic population into separate males and females. Interestingly, the gynoecium suppression function in S. latifolia evolved via loss of function in at least two sex-linked genes and works via gene dosage balance between sex-linked, and autosomal genes. This system resembles X/A-ratio-based sex determination systems in Drosophila and Rumex, and could represent a steppingstone in the evolution of X/A-ratio-based sex determination from an active Y system.

Postmeiotic irregular anther1 (PIA1) is required for anther cuticle and pollen exine development in maize (Zea mays L.)

AbstractMale sterile lines are valuable germplasm resources for hybrid seed production in maize (Zea mays L.). The structures of the anther cuticle and pollen exine are associated with male sterility. We obtained a completely male sterile mutant (postmeiotic irregular anther1, pia1), which has closed glumes and relatively small and wilted anthers (compared with the wild‐type control). Cytological analysis revealed the normal meiosis and premature tapetal degradation in pia1. Additionally, formation of the pia1 anther cuticle and Ubisch bodies was abnormal, and the pollen exine was discontinuous. Genetic analysis showed that the pia1 mutant was the result of a single recessive mutation of a nuclear gene. On the basis of fine mapping, PIA1 was mapped between the W07051 and W07124 molecular markers on chromosome 1. This region does not contain known genes associated with male sterility. Furthermore, transcriptome analysis indicated genes implicated in transmembrane transport, amino acid transmembrane transport, amino acid transport, and carboxylic acid transmembrane transport are differentially expressed in pia1 mutant anthers. The study findings suggest PIA1 is a novel regulator of anther cuticle and pollen exine development.

Rapid generation of a tomato male sterility system and its feasible application in hybrid seed production.

A practical approach for the rapid generation and feasible application of green hypocotyl male-sterile (GHMS) tm6 dfr lines in tomato hybrid breeding was established. Male sterility enables reduced cost and high seed purity during hybrid seed production. However, progress toward its commercial application has been slow in tomato due to the disadvantages of most natural male-sterile mutants. Here, we developed a practical method for efficient tomato hybrid seed production using a male-sterile system with visible marker, which was rapidly generated by CRISPR/Cas9-mediated gene editing. Two closely linked genes, TM6 and DFR, which were reported to be candidates of ms15 (male sterile-15) and aw (anthocyanin without) locus, respectively, were knocked out simultaneously in two elite tomato inbred lines. Mutagenesis of both genes generated green hypocotyl male-sterile (GHMS) lines. The GHMS lines exhibited male sterility across different genetic backgrounds and environmental conditions. They also showed green hypocotyl due to defective anthocyanin accumulation, which serves as a reliable visible marker for selecting male-sterile plants at the seedling stage. We further proposed a strategy for multiplying the GHMS system and verified its high efficiency in stable male sterility propagation. Moreover, elite hybrid seeds were produced using GHMS system for potential side effects evaluation, and no adverse influences were found on seed yield, seed quality as well as important agronomic traits. This study provides a practical approach for the rapid generation and feasible application of male sterility in tomato hybrid breeding.

Natural variation of MS2 confers male fertility and drives hybrid breeding in soybean.

A complete and genetically stable male sterile line with high outcrossing rate is a prerequisite for the development of commercial hybrid soybean. It was reported in the last century that the soybean male sterile ms2 mutant has the highest record with seed set. Here we report the cloning and characterization of the MS2 gene in soybean, which encodes a protein that is specifically expressed in the anther. MS2 functions in the tapetum and microspore by directly regulating genes involved in the biosynthesis of secondary metabolites and the lipid metabolism, which is essential for the formation of microspore cell wall. Through comparison of the field performance with the widely used male sterile mutants in the same genetic background, we demonstrated that the ms2 mutant conducts the best in outcrossing rate and makes it an ideal tool in building a cost-effective hybrid system for soybean.

OsTKPR1 proteins with a single amino acid substitution fail the synthesis of a specific sporopollenin precursor and cause abnormal exine and pollen development in rice

Fatty acid derivatives are key components of rice pollen exine. The synthesis of aliphatic sporopollenin precursors are initiated in the plastids of the tapetal cells, followed by multiple-step reactions conducted in the endoplasmic reticulum (ER). However, the relative contribution of different precursors to the precise structure of sporopollenin remains largely elusive, let alone the underlying mechanism. Here, we report that two complete male sterile mutants ostkpr1–3 (Tetraketide α-pyrone reductase 1–3, with OsTKPR1P124S substitution) and ostkpr1–4 (with truncated OsTKPR1stop) are defective in pollen exine, Ubisch body and anther cuticle development where ostkpr1–4 display severer phenotypes. Remarkably, OsTKPR1 could produce reduced hydroxylated tetraketide α-pyrone and reduced tetraketide α-pyrone, whereas OsTKPR1P124S fails to produce the latter. Pairwise interaction assays show that mutated OsTKPR1P124S is able to integrate into a recently characterized metabolon, thus its altered catalytic activity is not due to dis-integrity of the metabolon. In short, we find that reduced tetraketide α-pyrone is a key sporopollenin precursor required for normal exine formation, and the conserved 124th proline of OsTKPR1 is essential for the reduction activity. Therefore, this study provided new insights into the sporopollenin precursor constitution critical for exine formation.

Jasmonic acid-mediated stress responses share the molecular mechanism underlying male sterility induced by deficiency of ZmMs33 in maize

Plant male reproduction is a fine-tuned developmental process that is susceptible to stressful environments and influences crop grain yields. Phytohormone signaling functions in control of plant normal growth and development as well as in response to external stresses, but the interaction or crosstalk among phytohormone signaling, stress response, and male reproduction in plants remains poorly understood. Cross-species comparison among 514 stress-response transcriptomic libraries revealed that ms33-6038, a genic male sterile mutant deficient in the ZmMs33/ZmGPAT6 gene, displayed an excessive drought stress-like transcriptional reprogramming in anthers triggered mainly by disturbed jasmonic acid (JA) homeostasis. An increased level of JA appeared in ZmMs33-deficient anthers at both meiotic and post-meiotic stages and activated genes involved in JA biosynthesis and signaling as well as genes functioning in JA-mediated drought response. Excessive accumulation of JA elevated expression level of a gene encoding a WRKY transcription factor that activated the ZmMs33 promoter. These findings reveal a feedback loop of ZmMs33-JA-WRKY-ZmMs33 in controlling male sterility and JA-mediated stress response in maize, shedding light on the crosstalk of stress response and male sterility mediated by phytohormone homeostasis and signaling.

Induce male sterility by CRISPR/Cas9-mediated mitochondrial genome editing in tobacco.

Genome editing has become more and more popular in animal and plant systems following the emergence of CRISPR/Cas9 technology. However, target sequence modification by CRISPR/Cas9 has not been reported in the plant mitochondrial genome, mtDNA. In plants, a type of male sterility known as cytoplasmic male sterility (CMS) has been associated with certain mitochondrial genes, but few genes have been confirmed by direct mitochondrial gene-targeted modifications. Here, the CMS-associated gene (mtatp9) in tobacco was cleaved using mitoCRISPR/Cas9 with a mitochondrial localization signal. The male-sterile mutant, with aborted stamens, exhibited only 70% of the mtDNA copy number of the wild type and exhibited an altered percentage of heteroplasmic mtatp9 alleles; otherwise, the seed setting rate of the mutant flowers was zero. Transcriptomic analyses showed that glycolysis, tricarboxylic acid cycle metabolism and the oxidative phosphorylation pathway, which are all related to aerobic respiration, were inhibited in stamens of the male-sterile gene-edited mutant. In addition, overexpression of the synonymous mutations dsmtatp9 could restore fertility to the male-sterile mutant. Our results strongly suggest that mutation of mtatp9 causes CMS and that mitoCRISPR/Cas9 can be used to modify the mitochondrial genome of plants.