OsTDL1A binds to the LRR domain of rice receptor kinase MSP1, and is required to limit sporocyte numbers

Hybrids lose heterotic yield advantage when multiplied sexually via meiosis. A potential alternative breeding system for hybrids is apospory, where female gametes develop without meiosis. Common among grasses, apospory begins in the nucellus, where aposporous initials (AIs) appear near the sexual megaspore mother cell (MeMC). The cellular origin of AIs is obscure, but one possibility, suggested by the mac1 and msp1 mutants of maize and rice, is that AIs are apomeiotic derivatives of the additional MeMCs that appear when genetic control over sporocyte numbers is relaxed. MULTIPLE SPOROCYTES1 (MSP1) encodes a leucine-rich-repeat receptor kinase, which is orthologous to EXS/EMS1 in Arabidopsis. Like mac1 and msp1, exs/ems1 mutants produce extra sporocytes in the anther instead of a tapetum, causing male sterility. This phenotype is copied in mutants of TAPETUM DETERMINANT1 (TPD1), which encodes a small protein hypothesized to be an extracellular ligand of EXS/EMS1. Here we show that rice contains two TPD1-like genes, OsTDL1A and OsTDL1B. Both are co-expressed with MSP1 in anthers during meiosis, but only OsTDL1A and MSP1 are co-expressed in the ovule. OsTDL1A binds to the leucine-rich-repeat domain of MSP1 in yeast two-hybrid assays and bimolecular fluorescence complementation in onion cells; OsTDL1B lacks this capacity. When driven by the maize Ubiquitin1 promoter, RNA interference against OsTDL1A phenocopies msp1 in the ovule but not in the anther. Thus, RNAi produces multiple MeMCs without causing male sterility. We conclude that OsTDL1A binds MSP1 in order to limit sporocyte numbers. OsTDL1A-RNAi lines may be suitable starting points for achieving synthetic apospory in rice.     

PERSISTENT TAPETAL CELL1 encodes a PHD-finger protein that is required for tapetal cell death and pollen development in rice

In higher plants, timely degradation of tapetal cells, the innermost sporophytic cells of the anther wall layer, is a prerequisite for the development of viable pollen grains. However, relatively little is known about the mechanism underlying programmed tapetal cell development and degradation. Here, we report a key regulator in monocot rice (Oryza sativa), PERSISTANT TAPETAL CELL1 (PTC1), which controls programmed tapetal development and functional pollen formation. The evolutionary significance of PTC1 was revealed by partial genetic complementation of the homologous mutation MALE STERILITY1 (MS1) in the dicot Arabidopsis (Arabidopsis thaliana). PTC1 encodes a PHD-finger (for plant homeodomain) protein, which is expressed specifically in tapetal cells and microspores during anther development in stages 8 and 9, when the wild-type tapetal cells initiate a typical apoptosis-like cell death. Even though ptc1 mutants show phenotypic similarity to ms1 in a lack of tapetal DNA fragmentation, delayed tapetal degeneration, as well as abnormal pollen wall formation and aborted microspore development, the ptc1 mutant displays a previously unreported phenotype of uncontrolled tapetal proliferation and subsequent commencement of necrosis-like tapetal death. Microarray analysis indicated that 2,417 tapetum- and microspore-expressed genes, which are principally associated with tapetal development, degeneration, and pollen wall formation, had changed expression in ptc1 anthers. Moreover, the regulatory role of PTC1 in anther development was revealed by comparison with MS1 and other rice anther developmental regulators. These findings suggest a diversified and conserved switch of PTC1/MS1 in regulating programmed male reproductive development in both dicots and monocots, which provides new insights in plant anther development.     

ANTHER AND POLLEN DEVELOPMENT IN RICE (ORYZA SATIVA)

Investigations of the growth of anthers and ontogeny of pollen grains of Oryza sativa (rice) IR-30 were undertaken for the purpose of 1) providing a set of growth measurements and 2) describing stable cytological features of anther and pollen development. Correlations exist between elongation of the floret and growth parameters of the anther such as its length, width, fresh and dry weights and cytological stage of pollen development. In the early ontogeny of the anther, hypodermal archesporial initials divide periclinally to form primary parietal cells and primary sporogenous cells. Each of the latter divides twice mitotically to generate four microspore mother cells, which undergo meiosis. The anther wall is formed by anticlinal and periclinal divisions of the primary parietal cells as well as of cells surrounding the primary sporogenous cells. Subsequent cytological features in the development of anther and pollen grains of rice have much in common with anther and pollen developmental biology of other members of Gramineae.     

垂花悬铃花雌雄配子体发育研究

对垂花悬铃花雄配子体发育观察表明,其花药由表皮(1层)、药室内层(1层)、中层(2层)、绒毡层(1层)及造孢细胞组成,花药四室,药壁发育为双子叶型。雄配子体发育经由花粉母细胞减数分裂形成四分体,该四分体胞质分裂为同时型,四分体排列方式为四面体型,十字交叉型及左右对称型;小孢子再经有丝分裂形成营养核和生殖核,生殖核再经有丝分裂形成3-核花粉。花药壁层的变化,在单核小孢子期,表皮细胞解体,仅留下痕迹;中层在花粉母细胞期逐渐消失;药室内壁在单核小孢子期开始纤维化;绒毡层在单核小孢子期消失,属变形绒毡层。雌配子体发育观察表明,其子房上位,5室,每室1个胚珠,胚珠弯生,中轴胎座,大多数胚珠发育停留在珠心形成阶段,极少数珠心形成一群孢原细胞及单核、双核胚囊。     

The Post-meiotic Deficicent Anther1 （PDA1） gene is required for post-meiotic anther development in rice

To understand the molecular mechanism of male reproductive development in the model crop rice,we isolated a complete male sterile mutant post-meiotic deficient anther1 (pda1) from a γ-ray-treated rice mutant library.Genetic analysis revealed that the pda1 mutant was controlled by a recessive nucleus gene.The pda1 mutant anther seemed smaller with white appearance.Histological analysis demonstrated that the pda1 mutant anther undergoes normal early tapetum development without obvious altered meiosis.However,the pda1 mutant displayed obvious defects in postmeiotic tapetal development,abnormal degeneration occurred in the tapetal cells at stage 9 of anther development.Also we observed abnormal lipidic Ubisch bodies from the tapetal layer of the pda1 mutant,causing no obvious pollen exine formation.RT-PCR analysis indicated that the expression of genes involved in anther development including GAMYB,OsC4 and Wax-deficient anther1 (WDA1) was greatly reduced in the pda1 mutant anther.Using map-based cloning approach,the PDA1 gene was finely mapped between two markers HLF610 and HLF627 on chromosome 6 using 3,883 individuals of F2 population.The physical distance between HLF610 and HLF627 was about 194 kb.This work suggests that PDA1 is required for post-meiotic tapetal development and pollen/microspore formation in rice.     

The Cytological Mechanism of Low Fertility in the Naked Seed Rice

The low fertility of naked seed rice (NSR) was investigated by the following observations: somatic chromosome constitute, behavior of pollen mother cells (PMCs), the germination of mature pollen grains, the development of male and female gametes and the structure of the anther opening. The results indicated that somatic chromosomal number was 2n = 24, behavior of PMCs were normal and most of pollen grains could regularly develop further to mature male gametophytes in NSR. And dehiscence chamber and thickened endothecium cell (TEC) in numerous anthers of the NSR were developed abnormally after dicaryotic phase, result in few anthers complete opening and most partly opening or failure to opening, therefore much fewer of pollen grains attach on the stigma as compared with normal variety. Furthermore most of embryo sacs possessed abnormal structure and were sterile. All of above illustrated that the failure of the anther opening and the abortion of female gametophyte were main factors controlling the low seed-setting rate of the NSR.     

大花紫薇大小孢子的发生及雌雄配子体的发育

用石蜡切片法对不同发育时期的大花紫薇（Lagerstroemia speciosa）花朵进行解剖研究,探讨其大小孢子的发生及雌雄配子体的发育过程,结果发现：大花紫薇花药4室,花药壁由表皮、药室内壁、中层和腺质绒毡层构成,发育类型为双子叶型;小孢子四分体多为四面体型,偶见十字交叉型,胞质分裂为同时型;成熟花粉粒属于2-细胞型,具3孔沟,偶见败育现象;大花紫薇雌蕊具6~7心皮,子房6~7室,每室具多枚倒生胚珠,双珠被,厚珠心,大孢子4分体呈直线排列,近合点端大孢子发育为蓼型胚囊,成熟胚囊为7细胞8核。花粉及胚囊发育多数正常,大花紫薇可以作为优良的杂交母本;同时可以根据开花物候不同阶段花的形态特征,初步判断大花紫薇大、小孢子发生和雌、雄配子体的发育进程。     

RiceAntherNet: a gene co‐expression network for identifying anther and pollen development genes

In plants, normal anther and pollen development involves many important biological events and complex molecular regulatory coordination. Understanding gene regulatory relationships during male reproductive development is essential for fundamental biology and crop breeding. In this work, we developed a rice gene co‐expression network for anther development (RiceAntherNet) that allows prediction of gene regulatory relationships during pollen development. RiceAntherNet was generated from 57 rice anther tissue microarrays across all developmental stages. The microarray datasets from nine rice male sterile mutants, including msp1‐4, ostdl1a, gamyb‐2, tip2, udt1‐1, tdr, eat1‐1, ptc1 and mads3‐4, were used to explore and test the network. Among the changed genes, three clades showing differential expression patterns were constructed to identify genes associated with pollen formation. Many of these have known roles in pollen development, for example, seven genes in Clade 1 (OsABCG15, OsLAP5, OsLAP6, DPW, CYP703A3, OsNP1 and OsCP1) are involved in rice pollen wall formation. Furthermore, Clade 1 contained 12 genes whose predicted orthologs in Arabidopsis have been reported as key during pollen development and may play similar roles in rice. Genes in Clade 2 are expressed earlier than Clade 1 (anther stages 2–9), while genes in Clade 3 are expressed later (stages 10–12). RiceAntherNet serves as a valuable tool for identifying novel genes during plant anther and pollen development. A website is provided ( https://www.cpib.ac.uk/anther/riceindex.html ) to present the expression profiles for gene characterization. This will assist in determining the key relationships between genes, thus enabling characterization of critical genes associated with anther and pollen regulatory networks.     

濒危植物红花木莲小孢子发生及雄配子体发育的研究

首次报道了木莲属胚胎学特性,研究了红花木莲Manglietia insignis(Wall.)Bl.的小孢子发生、雄配子体的发育以及花粉萌发情况,红花木莲花药具有４个小孢子囊。花药壁５－６层,腺质绒毡层具有２－层细胞,小孢减数分裂后胞质分裂为修性同时型;四个分体排列方式为交叉型或左右对称型,偶为直线型;成熟花粉为二细胞型花粉粒,同时报道了化粉木莲在小孢子发生过程中的特有的现象,花粉萌发率为１４．７％－２５．３％。联系红花木莲花粉粒数目和胚珠数目进行探讨,认为雄配子体发育不是影响红花木莲结籽率低的主要因素。     

Two male-sterile mutants of Zea Mays (Poaceae) with an extra cell division in the anther wall

Two recessive male-sterile mutants of maize with similar patterns of pollen abortion were studied. Genetic studies showed that one of the two mutations was allelic with a previously identified male-sterility locus (ms23) and the other mutation was in a newly identified male-sterility locus (ms32). Cytological characterization of homozygous mutants and fertile heterozygous control siblings was performed using brightfield, fluorescence, and electron microscopy. During normal anther development, the final anther wall periclinal division divides the secondary parietal anther wall layer into the middle layer and tapetum, forming an anther with four wall layers. This is followed by differentiation of the tapetal cells into protoplastic binucleate, secretory tissue. In both the ms23 and ms32 mutants, the prospective tapetal layer divided into two layers, termed t1 and t2, forming an anther with five wall layers. Neither the t1 nor the t2 layers differentiated normally into tapetal layers, as determined by examination of cell walls, nucleus number, and cytoplasmic organization. Pollen mother cells aborted after the onset of prophase I of meiosis, suggesting that an early developmental coordination may exist between tapetum and pollen mother cells.     

柠条锦鸡儿小孢子发生和雄配子体发育

利用常规石蜡切片技术对柠条锦鸡儿小孢子发生及雄配子体发育的过程进行了观察,为柠条锦鸡儿生殖生物学提供基础资料。结果表明:(1)柠条锦鸡儿雄蕊花药4室,花药壁完全分化时,由外到内依次是表皮、药室内壁、中层和绒毡层,花药壁发育为基本型;表皮细胞1层,发育过程中始终存在;药室内壁在花药成熟时形成带状纤维层加厚;幼小花药壁的中层1～2层细胞,在花药发育成熟时退化消失;绒毡层1层细胞,腺质绒毡层,花药成熟时消失。(2)小孢子母细胞减数分裂过程中的胞质分裂为同时型,产生四面体型和左右对称型小孢子。(3)成熟花粉粒为二细胞型,扫描电镜下观察其成熟花粉粒为圆球形,外壁近光滑。(4)花粉母细胞分裂后形成的四分体小孢子中出现多核仁现象,核仁数在2～6个范围变化,推测这可能和末期Ⅱ核仁融合的不彻底有关。研究发现,柠条锦鸡儿小孢子发生和雄配子发育过程没有发现异常现象。     

华北驼绒藜大小孢子的发生及雌雄配子体发育过程的解剖学研究

运用石蜡切片技术对华北驼绒藜大、小孢子发生及雌、雄配子体的发育进行了研究.结果表明:(1)花药4室,花药壁由表皮、药室内壁、1~2层中层及1层绒毡层组成,药壁发育为基本型,腺质绒毡层,发育后期为二核或三核;(2)四分体胞质分裂为同时型,小孢子四分体多数为四面体型,偶见十字交叉型;(3)成熟花粉2-细胞型,单核小孢子时期存在不同比例的空壳花粉,从0%~80%不等;(4)胚珠倒生,双珠被,厚珠心,大孢子四分体直线型排列,合点端为功能大孢子,蓼型胚囊.     

The novel gene HOMOLOGOUS PAIRING ABERRATION IN RICE MEIOSIS1 of rice encodes a putative coiled-coil protein required for homologous chromosome pairing in meiosis

We have identified and characterized a novel gene, PAIR1 (HOMOLOGOUS PAIRING ABERRATION IN RICE MEIOSIS1), required for homologous chromosome pairing and cytokinesis in male and female meiocytes of rice (Oryza sativa). The pair1 mutation, tagged by the endogenous retrotransposon Tos17, exhibited meiosis-specific defects and resulted in complete sterility in male and female gametes. The PAIR1 gene encodes a 492-amino acid protein, which contains putative coiled-coil motifs in the middle, two basic regions at both termini, and a potential nuclear localization signal at the C terminus. Expression of the PAIR1 gene was detected in the early stages of flower development, in which the majority of the sporocytes had not entered meiosis. During prophase I of the pair1 meiocyte, all the chromosomes became entangled to form a compact sphere adhered to a nucleolus, and homologous pairing failed. At anaphase I and telophase I, chromosome nondisjunction and degenerated spindle formation resulted in multiple uneven spore production. However, chromosomal fragmentation frequent in plant meiotic mutants was never observed in all of the pair1 meiocytes. These observations clarify that the PAIR1 protein plays an essential role in establishment of homologous chromosome pairing in rice meiosis.     

关苍术两性花与雌花花药的解剖学研究

苍术属(Atractylodes DC.)是菊科菜蓟族(Cynareae)刺苞亚族(Carlininae O. Hoffm.)的一个东亚特有属,世界上仅有7种,其中我国有5种。该研究以关苍术为材料,采用石蜡切片法比较研究了两性花和雌花的花药及雄配子体发育进程,并进一步探讨了其雌花产生花药退化的时期及原因。结果表明：(1)关苍术小孢子发育与花蕾长度间存在相关性,当花蕾长度在5 mm时进入花粉母细胞时期,花药壁已分化,在7~9 mm时处于四分体时期,大于11 mm时开始进入花粉粒时期。(2)关苍术花药5个,花粉囊4个,减数分裂属同时型,四分体以正四面体为主,属3-细胞型,萌发沟3个。(3)关苍术花粉囊壁发育属双子叶型,从外层的表皮、药室内壁,到内层的中层和绒毡层均由一层细胞构成,关苍术绒毡层为腺质绒毡层。(4)关苍术雌花花药退化发生在花药发育早期至四分体时期,表现为花药发育早期畸形、药壁分化异常、小孢子母细胞发育停滞在前期、绒毡层增生4个原因。该研究结果为苍术属植物的系统发育、物种形成和进化提供胚胎学依据。     

矮牡丹小孢子发生和雄配子体发育及其与该种濒危的关系

研究了矮牡丹Paeonia jishanensis Hong et W．Z．Zhao的小孢子发生及雄配子体的形成。矮 牡丹花药具4个小孢子囊,药壁结构属双子叶型,腺质绒毡层,小孢子母细胞减数分裂后胞质分裂为 同时型,四分体多为四面体形,少左右对称形,成熟花粉为2-细胞。对芍药属木本类型的雄性发育进行 了全面研究,还对小孢子母细胞减数分裂和单核小孢子发育时期的异常现象进行了观察,对能育花粉 与不育花粉的百分比进行了测定,结果表明,能育花粉为45.03％～84.18％,它们在不同花中,不同花 药中,甚至同一花药的不同花粉囊中表现都不完全一致。联系矮牡丹的致濒原因进行了讨论,认为雄配子体形成过程中的异常现象,并不是导致矮牡丹濒危的主要因素。     

水稻花粉发育的分子机理

水稻的小孢子母细胞在花粉囊中进行减数分裂产生小孢子, 小孢子进一步发育成花粉粒。当花粉成熟时, 花粉粒从花粉囊中释放出来进行受精。分子生物学的研究已经发现了一些参与这一过程的基因, 包括控制花粉囊组织的分化、小孢子母细胞的减数分裂、小孢子的发育和花药的开裂等。本文旨在总结水稻花粉发育过程及其调控分子机制的研究进展。     

水稻花粉发育的分子机理

水稻的小孢子母细胞在花粉囊中进行减数分裂产生小孢子,小孢子进一步发育成花粉粒。当花粉成熟时,花粉粒从花粉囊中释放出来进行受精。分子生物学的研究已经发现了一些参与这一过程的基因,包括控制花粉囊组织的分化、小孢子母细胞的减数分裂、小孢子的发育和花药的开裂等。本文旨在总结水稻花粉发育过程及其调控分子机制的研究进展。     

山麦冬大小孢子发生及雌雄配子体发育研究

采用石蜡切片技术对百合科植物山麦冬大小孢子发生及雌雄配子体发育进行了观察研究。结果表明:(1)山麦冬花药具有4个花粉囊,花药壁的发育方式为基本型,花药壁完全分化时由表皮、药室内壁、中层及绒毡层组成。(2)绒毡层发育类型为分泌型,到四分体孢子彼此分离形成单细胞花粉阶段,绒毡层细胞开始解体退化,花粉成熟时绒毡层细胞完全消失;花粉母细胞减数分裂为连续型,四分体为左右对称形排列,成熟花粉为3-细胞花粉,单萌发沟。(3)子房3室,每室2枚胚珠,胚珠倒生型,双珠被,薄珠心,雌性孢原细胞不经过平周分裂而直接发育而成大孢子母细胞。(4)减数分裂后四分体大孢子呈线型或T型排列,合点端大孢子分化为功能大孢子,胚囊发育为蓼型;花粉母细胞减数分裂过程中,二分体、四分体细胞外方被胼胝质壁所包被,小孢子形成后胼胝质壁逐渐消失。该研究结果丰富了百合科植物生殖生物学研究的内容,也为探讨百合科植物的系统学研究提供了参考。