Anther Development Research Articles

Structure and development of anthers and connective glands in two species of Stigmaphyllon (Malpighiaceae): are heteromorphic anthers related to division of labour?

Heteranthery has been largely associated with a division of labour between anthers. Most species of Stigmaphyllon (Malpighiaceae) present heteromorphic anthers and glandular connectives of different development; yet, the functional meaning of this condition has never been explored in the genus. The aims of this study were to provide a comparative description of the structure and development of anthers and their connective glands in S. bonariense and S. jatrophifolium and to assess the existence of division of functions. Natural populations were selected to collect flowers at different stages. Anthers were subjected to morpho-anatomical, histochemical and pollen viability studies. For both species, abundance of pollen grains and size of anther and their connective glands were estimated. Three types of stamens are recognized: stamen with small, intermediate and large anthers. Anthers of both species exhibit a similar glandular tissue in the connective, and the histochemical analysis revealed that it produce a mucilagous secretion. The pattern of anther wall development, stainability and release of pollen grains was identical among anther types. For both species, we observed a positive relationship between anther size and abundance of pollen grains, but an inverse relationship between area of anthers and size (area and thickness) of connective glands in small anthers vs. intermediate and large ones. Our results evidence a specialization of anthers related to division of labour between heteromorphic stamens in two species of Stigmaphyllon. Thus, one set of anthers produces large amount of pollen grains for pollination and another sets large quantities of mucilage, which would improve pollen transport (better adherence to pollinator body and dampness maintenance). Nevertheless, heteranthery in both Stigmaphyllon species would represent a transitional state towards the division of labour rather than a stable state.

Expression pattern analysis of key genes related to anther development in a mutant of male-sterile Betula platyphylla Suk.

Male-sterile mutants are ideal materials for studying the gene regulatory pathway of anther development. Here, the cytological morphology of mutant and wild-type anthers during the whole development stage was analyzed. At the microspore developmental stage, the tapetum cells of two types of mutants (mutant male inflorescences (MM), normal-like male inflorescences (NLM) showed additional vacuoles and a reduced cytoplasm, while the wild-type tapetum cells began to degenerate. Before pollination, compared with formed mature pollen grains in wild-type male inflorescences, collapsed microspores were discovered in mutants. Based on the transcriptome data of wild-type male inflorescence (NM) and mutant MM, ten key differentially expressed genes (ABORTED MICROSPORES (AMS), MYB38, MALE STERILITY 1 (MS1), MYB32, ODORANT1 (ODO1), QUARTET3 (QRT3) and CYTOCHROME P450 77A3 (CYP77A3), ATP-Binding Cassette Transporter G26 (ABCG26), FIL1, LIPID-TRANSFER PROTEIN 14 (LTP14)) related to anther development were selected for further study. Furthermore, MYB32, MYB38, AMS, MS1, and ABCG26 were highly expressed at the tetrad stage in MM and NLM, along with ODO1, QRT3, and CYP77A3, which were significantly upregulated at the tetrad stage in NLM, FIL1 and LTP14 were upregulated at the tetrad stage in MM. These events suggested that these genes may accelerate the abortion process in mutant birch anther. Tissue-specific expression analysis of ten genes indicated diverse spatiotemporal expression patterns. QRT3 and CYP77A3 were expressed in all five tissues, including leaves, stems, roots, female inflorescences, and male inflorescences, while the other eight genes had high transcription levels during male development, indicating that these genes may play different roles in these developmental processes. These results facilitate further investigation of the functional and molecular mechanisms underlying anther and microspore development in Betula platyphylla.

Identification and profiling of microRNAs and differentially expressed genes during anther development between a genetic male-sterile mutant and its wildtype cotton via high-throughput RNA sequencing

Genetic male sterility (GMS) facilitates hybrid seed production in crops including cotton (Gossypium hirsutum). However, the genetic and molecular mechanisms specifically involved in this developmental process are poorly understood. In this study, small RNA sequencing, degradome sequencing, and transcriptome sequencing were performed to analyze miRNAs and their target genes during anther development in a GMS mutant (‘Dong A’) and its fertile wildtype (WT). A total of 80 known and 220 novel miRNAs were identified, 71 of which showed differential expressions during anther development. A further degradome sequencing revealed a total of 117 candidate target genes cleaved by 16 known and 36 novel miRNAs. Based on RNA-seq, 24, 11, and 21 predicted target genes showed expression correlations with the corresponding miRNAs at the meiosis, tetrad and uninucleate stages, respectively. In addition, a large number of differentially expressed genes were identified, most of which were involved in sucrose and starch metabolism, carbohydrate metabolism, and plant hormone signal transduction based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The results of our study provide valuable information for further functional investigations of the important miRNAs and target genes involved in genetic male sterility and advance our understanding of miRNA regulatory functions during cotton anther development.

OsMYB80 Regulates Anther Development and Pollen Fertility by Targeting Multiple Biological Pathways.

Pollen development is critical to the reproductive success of flowering plants, but how it is regulated is not well understood. Here, we isolated two allelic male-sterile mutants of OsMYB80 and investigated how OsMYB80 regulates male fertility in rice. OsMYB80 was barely expressed in tissues other than anthers, where it initiated the expression during meiosis, reached the peak at the tetrad-releasing stage and then quickly declined afterward. The osmyb80 mutants exhibited premature tapetum cell death, lack of Ubisch bodies, no exine and microspore degeneration. To understand how OsMYB80 regulates anther development, RNA-seq analysis was conducted to identify genes differentially regulated by OsMYB80 in rice anthers. In addition, DNA affinity purification sequencing (DAP-seq) analysis was performed to identify DNA fragments interacting with OsMYB80 in vitro. Overlap of the genes identified by RNA-seq and DAP-seq revealed 188 genes that were differentially regulated by OsMYB80 and also carried an OsMYB80-interacting DNA element in the promoter. Ten of these promoter elements were randomly selected for gel shift assay and yeast one-hybrid assay, and all showed OsMYB80 binding. The 10 promoters also showed OsMYB80-dependent induction when co-expressed in rice protoplast. Functional annotation of the 188 genes suggested that OsMYB80 regulates male fertility by directly targeting multiple biological processes. The identification of these genes significantly enriched the gene networks governing anther development and provided much new information for the understanding of pollen development and male fertility.

Lectin receptor kinase OsLecRK-S.7 is required for pollen development and male fertility.

Pollen grains are covered by exine that protects the pollen from stress and facilitates pollination. Here we isolated a male sterile mutant s13283 in rice exhibiting aborted pollen with abnormal exine and defective aperture. The mutant gene encodes a novel plasma membrane-localized legume-lectin receptor kinase that we named OsLecRK-S.7. OsLecRK-S.7 was expressed at different levels in all tested tissues and throughout anther development. In vitro kinase assay showed OsLecRK-S.7 capable of autophosporylation. Mutation in s13283 (E560K) and mutation of the conserved ATP binding site (K418E) both knocked out the kinase activity. Mass spectrometry showed Thr376 , Ser378 , Thr386 , Thr403 , and Thr657 to be the autophosphorylation sites. Mutation of individual autophosphorylation site affected the in vitro kinase activity to different degrees, but did not abolish the gene function in fertility complementation. oslecrk-s.7 mutant plant overexpressing OsLecRK-S.7 recovered male fertility but showed severe growth retardation with reduced number of tillers, and these phenotypes were abolished by E560K or K418E mutation. The results indicated that OsLecRK-S.7 was a key regulator of pollen development.

Impact of Autophagy on Gene Expression and Tapetal Programmed Cell Death During Pollen Development in Rice.

Autophagy has recently been shown to be required for tapetal programmed cell death (PCD) and pollen maturation in rice. A transcriptional regulatory network is also known to play a key role in the progression of tapetal PCD. However, the relationship between the gene regulatory network and autophagy in rice anther development is mostly unknown. Here, we comprehensively analyzed the effect of autophagy disruption on gene expression profile during the tapetal PCD in rice anther development using high-throughput RNA sequencing. Expression of thousands of genes, including specific transcription factors and several proteases required for tapetal degradation, fluctuated synchronously at specific stages during tapetal PCD progression in the wild-type anthers, while this fluctuation showed significant delay in the autophagy-deficient mutant Osatg7-1. Moreover, gene ontology enrichment analysis in combination with self-organizing map clustering as well as pathway analysis revealed that the expression patterns of a variety of organelle-related genes as well as genes involved in carbohydrate/lipid metabolism were affected in the Osatg7-1 mutant during pollen maturation. These results suggest that autophagy is required for proper regulation of gene expression and quality control of organelles and timely progression of tapetal PCD during rice pollen development.

Calcium distribution during anther development in Impatiens balsamina

The subcellular localization of loosely bound calcium in developing anthers of Impatiens balsamina (rose balzam) was investigated using potassium antimonate to precipitate the ions. At the sporogenous stage, few calcium precipitates were found in the young anthers. During meiosis of the microspore mother cell, the amount of calcium precipitates increased both inside these cells and in their cell walls. After meiosis, the resulting microspores accumulated abundant calcium precipitates in their cytoplasm and vacuoles. The microspores underwent mitosis to produce a bicellular pollen grain, in which the large vacuoles disintegrated, and the calcium precipitates of the vegetative cell were again localized in the cytoplasm. The amount of precipitates decreased during pollen maturity and finally disappeared from the inside of the pollen grains and accumulated on the pollen walls.

Morphological Features of the Anther Development in Tomato Plants with Non-Specific Male Sterility.

The study was devoted to morphological and cytoembryological analysis of disorders in the anther and pollen development of transgenic tomato plants with a normal and abnormal phenotype, which is characterized by the impaired development of generative organs. Various abnormalities in the structural organization of anthers and microspores were revealed. Such abnormalities in microspores lead to the blocking of asymmetric cell division and, accordingly, the male gametophyte formation. Some of the non-degenerated microspores accumulate a large number of storage inclusions, forming sterile mononuclear pseudo-pollen, which is similar in size and appearance to fertile pollen grain (looks like pollen grain). It was discussed that the growth of tapetal cells in abnormal anthers by increasing the size and ploidy level of nuclei contributes to this process. It has been shown that in transgenic plants with a normal phenotype, individual disturbances are also observed in the development of both male and female gametophytes. The reason for the developmental arrest of some ovules was the death of endosperm at different stages of the globular embryo. At the same time, noticeable hypertrophy of endothelial cells performing a secretory function was observed. In the ovules of transgenic plants with abnormalities, the endothelium forms a pseudo-embryo instead of the embryo sac, stimulating the development of parthenocarpic fruits. The data obtained in this study can be useful for a better understanding of the genetic and molecular mechanisms of cytoplasmic male sterility and parthenocarpic fruit development in tomatoes.

Different MicroRNA Families Involved in Regulating High Temperature Stress Response during Cotton (Gossypium hirsutum L.) Anther Development.

MicroRNAs (miRNAs) are small molecule RNAs widely involved in responses to plant abiotic stresses. We performed small RNA sequencing of cotton anthers at four developmental stages under normal and high temperature (NT and HT, respectively) conditions to investigate the stress response characteristics of miRNA to HT. A total of 77 miRNAs, including 33 known miRNAs and 44 novel miRNAs, were identified, and 41 and 28 miRNAs were differentially expressed under NT and HT stress conditions, respectively. The sporogenous cell proliferation (SCP), meiotic phase (MP), microspore release period (MRP), and pollen maturity (PM) stages had 10 (including 12 miRNAs), four (including six miRNAs), four (including five miRNAs), and seven (including 11 miRNAs) HT stress-responsive miRNA families, respectively, which were identified after removing the changes in genotype-specific miRNAs under NT condition. Seven miRNA families (miR2949, miR167, and miR160 at the SCP stage; miR156 and miR172 at the MP stage; miR156 at the MRP stage; and miR393 and miR3476 at the PM stage), which had expression abundance of more than 10% of the total expression abundance, served as the main regulators responding to HT stress with positive or negative regulation patterns. These miRNAs orchestrated the expression of the corresponding target genes and led to different responses in the HT-tolerant and the HT-sensitive lines. The results revealed that the HT stress response of miRNAs in cotton anthers were stage-specific and differed with the development of anthers. Our study may enhance the understanding of the response of miRNAs to HT stress in cotton anthers and may clarify the mechanism of plant tolerance to HT stress.

Transcriptome profiling of differentially expressed genes in cytoplasmic male-sterile line and its fertility restorer line in pigeon pea (Cajanus cajan L.)

BackgroundPigeon pea (Cajanus cajan L.) is the sixth major legume crop widely cultivated in the Indian sub-continent, Africa, and South-east Asia. Cytoplasmic male-sterility (CMS) is the incompetence of flowering plants to produce viable pollens during anther development. CMS has been extensively utilized for commercial hybrid seeds production in pigeon pea. However, the molecular basis governing CMS in pigeon pea remains unclear and undetermined. In this study transcriptome analysis for exploring differentially expressed genes (DEGs) between cytoplasmic male-sterile line (AKCMS11) and its fertility restorer line (AKPR303) was performed using Illumina paired-end sequencing.ResultsA total of 3167 DEGs were identified, of which 1432 were up-regulated and 1390 were down-regulated in AKCMS11 in comparison to AKPR303. By querying, all the 3167 DEGs against TAIR database, 34 pigeon pea homologous genes were identified, few involved in pollen development (EMS1, MS1, ARF17) and encoding MYB and bHLH transcription factors with lower expression in the sterile buds, implying their possible role in pollen sterility. Many of these DEGs implicated in carbon metabolism, tricarboxylic acid cycle (TCA), oxidative phosphorylation and elimination of reactive oxygen species (ROS) showed reduced expression in the AKCMS11 (sterile) buds.ConclusionThe comparative transcriptome findings suggest the potential role of these DEGs in pollen development or abortion, pointing towards their involvement in cytoplasmic male-sterility in pigeon pea. The candidate DEGs identified in this investigation will be highly significant for further research, as they could lend a comprehensive basis in unravelling the molecular mechanism governing CMS in pigeon pea.

Sublethal application of various sulfonylurea and imidazolinone herbicides favors outcrossing and hybrid seed production in oilseed rape

BackgroundAcetolactate synthase (ALS)-inhibiting herbicides from the chemical families of sulfonylureas and imidazolinones are used worldwide. However, drift or sprayer contamination from some sulfonylurea herbicides causes a high level of male sterility in cruciferous species, especially oilseed rape (OSR). In this paper, we evaluated the gametocidal effects of 27 ALS-inhibiting herbicides that were sprayed on OSR plants at the bolting stage.ResultsOSR anther development was very sensitive to sublethal exposure to most ALS-inhibiting herbicides. The application of 18 out of the 20 tested sulfonylureas (except ethametsulfuron and ethoxysulfuron), two imidazolinones (imazethapyr and imazamox), and one sulfonylamino-carbonyltriazolinone (flucarbazone-sodium) at suitable rates could induce male sterility. Eight of the herbicides, including chlorsulfuron (at application rates of 60–120 mg/ha), halosulfuron-methyl (300–600 mg/ha), sulfosulfuron (400–600 mg/ha), triflusulfuron-methyl (500–750 mg/ha), pyrazosulfuron-ethyl (150–225 mg/ha), nicosulfuron (200–300 mg/ha), imazethapyr (750–1125 mg/ha), and imazamox (400–800 mg/ha), could induce over 90% male sterility and over 60% relative outcrossed seed set in six cultivars with different origins. These eight chemicals could be used as new gametocides for hybrid seed production. This study also examined the possibility of external application of these gametocides on several unstable Polima cytoplasmic male sterile and thermosensitive genic male sterile lines. Although the outcrossed seed set of the treated lines was slightly reduced, the gametocide application significantly increased the seed purity of the resulting hybrid.ConclusionThe finding of the gametocidal effects of most sulfonylureas and imidazolinones are of great importance for developing new functions for ALS-inhibiting herbicides. The application of gametocides will also greatly promote the safe utilization of environment-sensitive male sterility in hybrid seed production. Unexpectedly, the application of three triazolopyrimidines (florasulam, flumetsulam, and penoxsulam) and one pyrimidinylthiobenzoate (bispyribac-sodium) did not cause male sterility, although these herbicides obviously inhibited the activity of ALS and plant growth. This result suggests that inhibition of ALS activity does not always lead to male sterility in plants, and these gametocides may also inhibit other biological functions vital for microspore development.

Monitoring rice anther proteome expression patterns during pollen development

Rice (Oryza sativa L.) pollen grains are highly sensitive to extreme temperatures. Studies on the proteomic analysis of pollen at different developmental stages are limited. In this study, to identify proteins expressed during pollen development, we performed shotgun proteomic analysis of rice anthers harvested at 8, 3, and 0 days before heading (DBH). A total of 4509 non-redundant proteins were identified in rice anthers at each of the three time points with three replications per time point. Among these, 2968 proteins were reproducible, of which 1919 were specifically expressed at a given stage of anther development. Clustering and gene ontology (GO) enrichment analyses categorized the specifically expressed proteins into four groups. Group I comprised proteins involved in cell differentiation, reproduction, and stress response; Group II contained proteins related to cell growth and carbohydrate metabolic process; Group III included proteins related to starch accumulation and lipid metabolic process; Group IV comprised proteins related to structure morphogenesis and cellular components. Proteins in Group I steadily increased during anther development, whereas those in Group I steadily decreased. Group III proteins showed the highest expression at 3 DBH, whereas Group IV proteins exhibited the lowest expression at 3 DBH.

RMS2 Encoding a GDSL Lipase Mediates Lipid Homeostasis in Anthers to Determine Rice Male Fertility.

Plant male gametogenesis is a coordinated effort involving both reproductive tissues and sporophytic tissues, in which lipid metabolism plays an essential role. Although GDSL esterases/lipases have been well known as key enzymes for many plant developmental processes and stress responses, their functions in reproductive development remain unclear. Here, we report the identification of a rice male sterile2 (rms2) mutant in rice (Oryza sativa), which is completely male sterile due to the defects in tapetum degradation, cuticle formation in sporophytic tissues, and impaired exine and central vacuole development in pollen grains. RMS2 was map-based cloned as an endoplasmic reticulum-localized GDSL lipase gene, which is predominantly transcribed during early anther development. In rms2, a three-nucleotide deletion and one base substitution (TTGT to A) occurred within the GDSL domain, which reduced the lipid hydrolase activity of the resulting protein and led to significant changes in the content of 16 lipid components and numerous other metabolites, as revealed by a comparative metabolic analysis. Furthermore, RMS2 is directly targeted by the male fertility regulators Undeveloped Tapetum1 and Persistent Tapetal Cell1 both in vitro and in vivo, suggesting that RMS2 may serve as a key node in the rice male fertility regulatory network. These findings shed light on the function of GDSLs in reproductive development and provide a promising gene resource for hybrid rice breeding.

ERdj3B-Mediated Quality Control Maintains Anther Development at High Temperatures.

Pollen development is highly sensitive to heat stress, which impairs cellular proteostasis by causing misfolded proteins to accumulate. Therefore, each cellular compartment possesses a dedicated protein quality control system. An elaborate quality control system involving molecular chaperones, including immunoglobulin-binding protein (BiP), heat shock protein70, and regulatory J domain-containing cochaperones (J proteins), allows the endoplasmic reticulum (ER) to withstand a large influx of proteins. Here, we found that Arabidopsis (Arabidopsis thaliana) mutants of ER-localized DnaJ family 3B (ERdj3B), one of three ER-resident J proteins involved in ER quality control, produced few seeds at high temperatures (29°C) due to defects in anther development. This temperature-sensitive fertility defect is specific to the defective interactions of BiP with ERdj3B but not with the other two J proteins, indicating functional differences between ERdj3B and the other J proteins. RNA sequencing analysis revealed that heat stress affects pollen development in both wild-type and mutant buds, but the erdj3b mutant is more susceptible, possibly due to defects in ER quality control. Our results highlight the importance of a specific ER quality control factor, ERdj3B, for plant reproduction, particularly anther development, at high temperatures.

Acute heat stress during stamen development affects both the germline and sporophytic lineages in Arabidopsis thaliana (L.) Heynh.

Flowers, from their initiation in the inflorescence meristem until their opening at anthesis, are particularly sensitive to subtle changes in environmental conditions. Short episodes of either cold or heat stress experienced during stamen development may lead to varying degrees of male sterility and can affect the ploidy of the gametes. In an attempt to understand the developmental origins of these and other phenotypes that are caused by harsh environmental conditions, an acute heat stress was simultaneously applied to all stages of anther development, and its consequences on pollen viability at anthesis and seed set were traced back to heat stress at specific developmental stages. We identified a critical stage during microsporogenesis that is highly sensitive to heat stress. The acute heat stress experienced during these stages completely blocked stamen and pollen development, likely mediated by its effects on both anther wall development and the formation of polyads in the germline. Interestingly, as we investigated heat shocks at earlier developmental stages, a gradual recovery of male fertility was observed, characterized by some abnormalities in locule development and by fusions of maturing pollen grains. Overall, our findings establish a developmental framework for the diverse effects of heat stress on stamen and pollen development, which is relevant to different fields of research. Acute heat stresses are expected to increase in frequency with global warming and might negatively affect plant fitness and crop yields in the short term by affecting male fertility and seed set. Moreover, the formation of polyads and the fusion of pollen grains offer insights into the developmental origins of evolutionary relevant processes, such as polyploidization, which might offer opportunities for environmental adaptations in the long term.

Transcriptional trajectories of anther development provide candidates for engineering male fertility in sorghum

Sorghum is a self-pollinated crop with multiple economic uses as cereal, forage, and biofuel feedstock. Hybrid breeding is a cornerstone for sorghum improvement strategies that currently relies on cytoplasmic male sterile lines. To engineer genic male sterility, it is imperative to examine the genetic components regulating anther/pollen development in sorghum. To this end, we have performed transcriptomic analysis from three temporal stages of developing anthers that correspond to meiotic, microspore and mature pollen stages. A total of 5286 genes were differentially regulated among the three anther stages with 890 of them exhibiting anther-preferential expression. Differentially expressed genes could be clubbed into seven distinct developmental trajectories using K-means clustering. Pathway mapping revealed that genes involved in cell cycle, DNA repair, regulation of transcription, brassinosteroid and auxin biosynthesis/signalling exhibit peak expression in meiotic anthers, while those regulating abiotic stress, carbohydrate metabolism, and transport were enriched in microspore stage. Conversely, genes associated with protein degradation, post-translational modifications, cell wall biosynthesis/modifications, abscisic acid, ethylene, cytokinin and jasmonic acid biosynthesis/signalling were highly expressed in mature pollen stage. High concurrence in transcriptional dynamics and cis-regulatory elements of differentially expressed genes in rice and sorghum confirmed conserved developmental pathways regulating anther development across species. Comprehensive literature survey in conjunction with orthology analysis and anther-preferential accumulation enabled shortlisting of 21 prospective candidates for in-depth characterization and engineering male fertility in sorghum.

Genome-Wide Investigation of Heat Shock Transcription Factor Family in Wheat (Triticum aestivum L.) and Possible Roles in Anther Development.

Heat shock transcription factors (HSFs) play crucial roles in resisting heat stress and regulating plant development. Recently, HSFs have been shown to play roles in anther development. Thus, investigating the HSF family members and identifying their protective roles in anthers are essential for the further development of male sterile wheat breeding. In the present study, 61 wheat HSF genes (TaHsfs) were identified in the whole wheat genome and they are unequally distributed on 21 chromosomes. According to gene structure and phylogenetic analyses, the 61 TaHsfs were classified into three categories and 12 subclasses. Genome-wide duplication was identified as the main source of the expansion of the wheat HSF gene family based on 14 pairs of homeologous triplets, whereas only a very small number of TaHsfs were derived by segmental duplication and tandem duplication. Heat shock protein 90 (HSP90), HSP70, and another class of chaperone protein called htpG were identified as proteins that interact with wheat HSFs. RNA-seq analysis indicated that TaHsfs have obvious period- and tissue-specific expression patterns, and the TaHsfs in classes A and B respond to heat shock, whereas the C class TaHsfs are involved in drought regulation. qRT-PCR identified three TaHsfA2bs with differential expression in sterile and fertile anthers, and they may be candidate genes involved in anther development. This comprehensive analysis provides novel insights into TaHsfs, and it will be useful for understanding the mechanism of plant fertility conversion.

Cytological atlas at meiosis reveals insights into pollen fertility in synthetic Brassica allotriploids between allotetraploid B. carinata and diploid B. rapa

The formation of allopolyploid crops basically depends on the successful interspecific hybridization and polyploidization, which generally involves in a combination of distinct but related genomes from independent species. But cytological analysis of these initially synthesized allohaploids immediately after genome merging is poorly explored in regards to anther and pollen development to date. In this study, Brassica trigenomic allohaploids (ABC) were produced to investigate the immediate effects of the genome combinations on pollen fertility during anther development via crosses between natural allotetraploid B. carinata (BBCC) and diploid B. rapa (AA). The results showed that in the synthetic Brassica allotriploids (ABC), the anther development was completely disrupted, and the pollen grains were mostly inviable with varied genetic complements. In addition, the meiosis course was aberrantly altered and eccentric chromosomal configurations including multivalent, bridges and lags occurred frequently during metaphase I to anaphase II. Genomic in situ hybridization (GISH) further revealed that B genome of homoeology was frequently apt to interact with A and C genomes, and cytoskeletal organizations was improperly distributed during meiosis in these synthetic Brassica allotriploids. Furthermore, we also confirmed that the expression of typical meiosis-related genes was obviously repressed during anther development in these Brassica allotriploids. Taken together, our results provide a detailed cytology for insights into pollen development in the synthetic allotriploid hybrids, which are conventionally considered as a useful genetic resource for polyploid Brassica breeding.

Cytological observation of anther structure and genetic investigation of a thermo-sensitive genic male sterile line 373S in Brassica napus L

BackgroundPhotoperiod and/or thermo-sensitive male sterility is an effective pollination control system in crop two-line hybrid breeding. We previously discovered the spontaneous mutation of a partially male sterile plant and developed a thermo-sensitive genic male sterile (TGMS) line 373S in Brassica napus L. The present study characterized this TGMS line through cytological observation, photoperiod/ temperature treatments, and genetic investigation.ResultsMicroscopic observation revealed that the condensed cytoplasm and irregular exine of microspores and the abnormal degradation of tapetum are related to pollen abortion. Different temperature and photoperiod treatments in field and growth cabinet conditions indicated that the fertility alteration of 373S was mainly caused by temperature changes. The effects of photoperiod and interaction between temperature and photoperiod were insignificant. The critical temperature leading to fertility alteration ranged from 10 °C (15 °C/5 °C) to 12 °C (17 °C/7 °C), and the temperature-responding stage was coincident with anther development from pollen mother cell formation to meiosis stages. Genetic analysis indicated that the TGMS trait in 373S was controlled by one pair of genes, with male sterility as the recessive. Multiplex PCR analysis revealed that the cytoplasm of 373S is pol type.ConclusionsOur study suggested that the 373S line in B. napus has a novel thermo-sensitive gene Bnmst1 in Pol CMS cytoplasm background, and its fertility alteration is mainly caused by temperature changes. Our results will broaden the TGMS resources and lay the foundation for two-line hybrid breeding in B. napus.

Identification and Analysis of <i>R2R3-MYB</i> Genes in Sweet Potato Genome

The plant MYB is a transcription factor family large in number and with important functions. In this study, the MYB family genes were screened and identified via bioinformatic methods from the raw sequence of sweet potato ( Ipomoea batatas ) genome and the gene structure and function of R2R3-MYB were analyzed. The results showed that there were 88 R2R3-MYB genes with intact R2 and R3 conservative domains, which contained 8 and 9 highly conserved basic amino acids. The results of MEME analysis showed that there were 10 conserved motifs within the I. batatas R2R3-MYB protein sequences. For the I. batatas R2R3-MYB protein sequences, over 80% contained motif 1, motif 2, motif 3, motif 4, motif 5 and motif 7. The R2R3-MYB genes were distributed unevenly across the 15 chromosomes. The number of R2R3-MYB genes in chromosome No.5 was 15, which was the largest; the numbers of R2R3-MYB genes in chromosome No. 4 and 13 were both only 2, which was the smallest. Analysis of the sequence alignment showed that there were 6 pairs of interchromosomal duplication and there were 20 pairs of intrachromosomal duplication, 19 of which existed in clusters. The function prediction and categorization via sequence analysis showed that 44 R2R3-MYB genes of the I. batatas could be categorized to the 13 subgroups of the A. thaliana R2R3-MYB genes, which were involved in the responses to biotic stress and abiotic stress, anthocyanin biosynthesis, anther development, etc. Further analysis showed that 36 R2R3-MYB genes might play important roles in dealing with biotic stress and abiotic stress, 9 of which showed significant up/down-regulation under Fusarium oxysporum f. sp. batatas stress and 27 of which showed significant up/down-regulation under low temperature stress. The domains of I. batatas R2R3-MYB transcription factors were highly conservative, which contained highly conserved motifs within R2 and R3 domains. The phylogenetic tree and transcriptomics data analysis showed that some R2R3-MYB genes might play roles in growth and development, metabolism regulation, biotic stress and abiotic stress, which lent support to I. batatas breeding.