Tapetal Cells Research Articles

Melatonin Alleviates High Temperature-Induced Pollen Abortion in Solanum lycopersicum.

Melatonin is a pleiotropic signal molecule that plays critical roles in regulating plant growth and development, as well as providing physiological protections against various environmental stresses. Nonetheless, the mechanisms for melatonin-mediated pollen thermotolerance remain largely unknown. In this study, we report that irrigation treatment with melatonin (20 µM) effectively ameliorated high temperature-induced inactivation of pollen and inhibition of pollen germination in tomato (Solanum lycopersicum) plants. Melatonin alleviated reactive oxygen species production in tomato anthers under high temperature by the up-regulation of the transcription and activities of several antioxidant enzymes. Transmission electron micrograph results showed that high temperature-induced pollen abortion is associated with a premature degeneration of the tapetum cells and the formation of defective pollen grains with degenerated nuclei at the early uninuclear microspore stage, whilst melatonin protected degradation of organelles by enhancing the expression of heat shock protein genes to refold unfolded proteins and the expression of autophagy-related genes and formation of autophagosomes to degrade denatured proteins. These findings suggest a novel function of melatonin to protect pollen activity under high temperature and support the potential effects of melatonin on reproductive development of plants.

OsPKS2 is required for rice male fertility by participating in pollen wall formation.

OsPKS2, the rice orthologousgene of Arabidopsis PKSB/LAP5, encodes a polyketide synthase that is involved in pollen wall formation in rice. In flowering plants, the pollen wall protects male gametes from various environmental stresses and pathogen attacks, as well as promotes pollen germination. The biosynthesis of sporopollenin in tapetal cell is critical for pollen wall formation. Recently, progress has been made in understanding sporopollenin metabolism during pollen wall development in Arabidopsis. However, little is known about the molecular mechanism that underlies the sporopollenin synthesis in pollen wall formation in rice (Oryza sativa). In this study, we identified that a point mutation in OsPKS2, a plant-specific type III polyketide synthase gene, caused male sterility in rice by affecting the normal progress of pollen wall formation. Two other allelic mutants of OsPKS2 were generated using the CRISPR/Cas9 system and are also completely male sterile. This result thus further confirmed that OsPKS2 controls rice male fertility. We also showed that OsPKS2 is an orthologous gene of Arabidopsis PKSB/LAP5 and has a tapetum-specific expression pattern. In addition, its product localizes in the endoplasmic reticulum. Results suggested that OsPKS2 is critical for pollen wall formation, and plays a conserved but differentiated role in sporopollenin biosynthesis from Arabidopsis.

Identification of cyp703a3-3 and analysis of regulatory role of CYP703A3 in rice anther cuticle and pollen exine development

Anther cuticle and pollen exine are two elaborated lipid-soluble barriers protecting pollen grains from environmental and biological stresses. However, less is known about the mechanisms underlying the synthesis of these lipidic polymers. Here, we identified a no-pollen male-sterility mutant cyp703a3-3 from the indica restorer line Zhonghui 8015 (Zh8015) mutant library treated with 60Coγ-ray radiation. Histological analysis indicated that cyp703a3-3 underwent abnormal tapetal cells development, produced few orbicules and secreted less sporopollenin precursors to anther locule, as well as cutin monomers on anther. Genetic analysis revealed that cyp703a3-3 was controlled by a single recessive gene. Map-based cloning was performed to narrow down the mutant gene to a 47.78-kb interval on the chromosome 8 between two markers S15-29 and S15-30. Sequence analysis detected three bases (GAA) deletion in the first exon of LOC_Os08g03682, annotated as CYP703A3 with homologous sequences related to male sterility in Arabidopsis, causing the Asparagine deletion in the mutant site. Moreover, we transformed genomic fragment of CYP703A3 into cyp703a3-3, which male-sterility phenotype was recovered. Both the wild-type and cyp703a3-3 mutant 3D structure of CYP703A3 protein were modeled. Results of qPCR suggested CYP703A3 mainly expressed in anthers with greatest abundance at microspore stage, and genes involved in sporopollenin precursors formation and transportation, such as GAMYB, TDR, CYP704B2, DPW2, OsABCG26 and OsABCG15, were significantly reduced in cyp703a3-3. Collectively, our results further elaborated CYP703A3 plays vital role in anther cuticle and pollen exine development in rice (Oryza sativa L.).

The Arabidopsis COPII components, AtSEC23A and AtSEC23D, are essential for pollen wall development and exine patterning.

The specialized multilayered pollen wall plays multiple roles to ensure normal microspore development. The major components of the pollen wall (e.g. sporopollenin and lipidic precursors) are provided from the tapetum. Material export from the endoplasmic reticulum (ER) is mediated by coat protein complex II (COPII) vesicles. The Arabidopsis thaliana genome encodes seven homologs of SEC23, a COPII component. However, the functional importance of this diversity remains elusive. Here, we analyzed knockout and knockdown lines for AtSEC23A and AtSEC23D, two of the A. thaliana SEC23 homologs, respectively. Single atsec23a and atsec23d mutant plants, despite normal fertility, showed an impaired exine pattern. Double atsec23ad mutant plants were semi-sterile and exhibited developmental defects in pollen and tapetal cells. Pollen grains of atsec23ad had defective exine and intine, and showed signs of cell degeneration. Moreover, the development of tapetal cells was altered, with structural abnormalities in organelles. AtSEC23A and AtSEC23D exhibited the characteristic localization pattern of COPII proteins and were highly expressed in the tapetum. Our work suggests that AtSEC23A and AtSEC23D may organize pollen wall development and exine patterning by regulating ER export of lipids and proteins necessary for pollen wall formation. Also, our results shed light on the functional heterogeneity of SEC23 homologs.

Barnase-barstar system: an indelible technique to produce hybrid seeds in self-pollinated crops

Self-pollinated crops are sensitive to exploit high level of heterosis. The improvement of crop plants through the production of hybrid varieties is a major goal of plant breeding. The genetically engineered techniques have a pivotal role in developing male sterility and restorer systems. Chimaeric ribonuclease gene (barnase) expression within the anther selectively destroys the tapetal cell layer that surrounds the pollen sac, prevents pollen formation and leads to male sterility. A chimaeric tapetal cell-specific ribonuclease inhibitor gene (barstar) was used in male fertile plants to restore male fertility in F1 progeny. Cytotoxic ribonuclease activity in the anther was suppressed to restore male fertility in F1 hybrid by the formation of cell-specific RNase/RNase inhibitor complexes.

Identification of Anther-Specific Genes from Sesame and Functional Assessment of the Upstream Region of a Tapetum-Specific β-1,3-glucanase Gene

Sesamum indicum is widely cultivated in India for its superior agronomic characters like adaptation to the warm climate of India and resistance to drought and pathogen attack. However, the yield of sesame per unit area of land is quite low. In this context, the genes involved with reproductive tissue development is very important since they are directly involved with crop productivity. Therefore, identification and isolation of anther-specific genes through PCR-Select cDNA subtraction between cDNAs of anthers and leaves was carried out. Sequencing of ~ 700 clones from the cDNA subtraction library followed by BLASTX analysis using sesame database resulted in the identification of 163 annotated unique transcripts, of which 18 transcripts showed homology with the genes of exclusive anther-/pollen-specific functions. Furthermore, anther-specific expression of some of the sesame transcripts as well as already identified anther-specific genes of Arabidopsis and rice have been monitored during development of sesame anthers. Subsequently, full-length cDNA sequence and the upstream sequence of an anther-specific β-1 3-glucanase gene was isolated. Transgenic tobacco plants were developed with GUS gene under the control of a 437-bp upstream sequence along with the first exon and first intron of the gene. Monitoring of GUS gene expression pattern indicated exclusive expression of the promoter in the tapetal cell layer before tetrad formation in the anther. Interestingly, upon treatment with ABA as well as drought stress, GUS was found to express in the vegetative parts of the plant such as leaves and roots as well.

Fine regulation of ARF17 for anther development and pollen formation

BackgroundIn Arabidopsis, the tapetum and microsporocytes are critical for pollen formation. Previous studies have shown that ARF17 is expressed in microsporocytes and tetrads and directly regulates tetrad wall synthesis for pollen formation. ARF17 is the direct target of miR160, and promoterARF17::5mARF17 (5mARF17/WT) transgenic plants, which have five silent mutations within the miR160-complementary domain, are sterile.ResultsHere, we found that ARF17 is also expressed in the tapetum, which was defective in arf17 mutants. Compared with arf17 mutants, 5mARF17/WT plants had abnormal tapetal cells and tetrads but were less vacuolated in the tapetum. Immunocytochemical assays showed that the ARF17 protein over-accumulated in tapetum, microsporocytes and tetrads of 5mARF17/WT plants at early anther stages, but its expression pattern was not affected during anther development. 5mARF17 driven by its native promoter did not rescue the arf17 male-sterile phenotype. The expression of 5mARF17 driven by the tapetum-specific promoter A9 led to a defective tapetum and male sterility in transgenic plants. These results suggest that the overexpression of ARF17 in the tapetum and microsporocytes of 5mARF17/WT plants leads to male sterility. Microarray data revealed that an abundance of genes involved in transcription and translation are ectopically expressed in 5mARF17/WT plants.ConclusionsOur work shows that ARF17 plays an essential role in anther development and pollen formation, and ARF17 expression under miR160 regulation is critical for its function during anther development.

ABA and IAA control microsporogenesis in Petunia hybrida L.

The formation of fertile male gametophyte is known to require timely degeneration of polyfunctional tapetum tissue. The last process caused by the programmed cell death (PCD) is a part of the anther program maturation which leads to sequential anther tissue destruction coordinated with pollen differentiation. In the present work, distribution of abscisic acid (ABA) and indole-3-acetic acid (IAA) in developing anthers of male-fertile and male-sterile lines of petunia (Petunia hybrida L.) was analyzed by using the immunohistochemical method. It was established that the development of fertile male gametophyte was accompanied by monotonous elevation of ABA and IAA levels in reproductive cells and, in contrast, their monotonous lowering in tapetum cells and the middle layers. Abortion of microsporocytes in the meiosis prophase in the sterile line caused by premature tapetum degeneration along with complete maintenance of the middle layers was accompanied by dramatic, twofold elevation in the levels of both the phytohormones in reproductive cells. The data obtained allowed us to conclude that at the meiosis stage ABA and IAA are involved in the PCD of microsporocytes.

Ultrastructural abnormalities in pollen and anther wall development may lead to low pollen viability in jasmine (Jasminum sambac (L.) Aiton, Oleaceae)

Jasmine [Jasminum sambac (L.) Aiton] rarely sets seed, and its poor pollen fertility significantly influences fecundity. In this study, pollen development of jasmine was investigated at an ultrastructural level, examining the dynamic changes of microspore-tetrads and tapetum and their possible effects on pollen viability. Normally developed microspores are released at the late tetrad phase, whereas the abnormal ones formed large complexes of two to five tetrads with plasmodesmata. Ultrastructural observation also revealed that sterility of some pollen grains might result from defective development of cell walls that failed to absorb their surrounding sporopollenin to form the exine. The secretory type of jasmine tapetum, the usually degenerated middle layer and thickened endothecium were confirmed during the process of pollen development. The epidermis, middle layer and endothecium exhibited no adverse effects on pollen development and dispersal. However, the asynchronous division and delayed degradation of tapetal cells resulted in abnormal development of some microspores and the formation of massulae and empty and flat pollen grains. This is the first ultrastructural characterization of pollen development in J. sambac leading to a better understanding of embryology and poor pollen fertility of this important horticultural species.

Evolvement of transgenic male-sterility and fertility-restoration system in rice for production of hybrid varieties.

We have developed a unique male-sterility and fertility-restoration system in rice by combining Brassica napus cysteine-protease gene (BnCysP1) with anther-specific P12 promoter of rice for facilitating production of hybrid varieties. In diverse crop plants, male-sterility has been exploited as a useful approach for production of hybrid varieties to harness the benefits of hybrid vigour. The promoter region of Os12bglu38 gene of rice has been isolated from the developing panicles and was designated as P12. The promoter was fused with gusA reporter gene and was expressed in Arabidopsis and rice systems. Transgenic plants exhibited GUS activity in tapetal cells and pollen of the developing anthers indicating anther/pollen-specific expression of the promoter. For engineering nuclear male sterility, the coding region of Brassica napus cysteine protease1 (BnCysP1) was isolated from developing seeds and fused to P12 promoter. Transgenic rice plants obtained with P12-BnCysP1 failed to produce functional pollen grains. The F1 seeds obtained from BnCysP1 male-sterile plants and untransformed controls showed 1:1 (tolerant:sensitive) ratio when germinated on the MS medium supplemented with phosphinothricin (5mg/l), confirming that the male sterility has been successfully engineered in rice. For male fertility restoration, transgenic rice plants carrying BnCysP1Si silencing system were developed. The pollination of BnCysP1 male-sterile (female-fertile) plants with BnCysP1Si pollen resulted in normal grain filling. The F1 seeds of BnCysP1 × BnCysP1Si when germinated on the MS basal medium containing PPT (5mg/l) and hygromycin (70mg/l) exhibited 1:1 (tolerant:sensitive) ratio and the tolerant plants invariably showed normal grain filling. The overall results clearly suggest that the customized male-sterility & fertility-restoration system can be exploited for quality hybrid seed production in various crops.

Two rice receptor-like kinases maintain male fertility under changing temperatures

Plants employ dynamic molecular networks to control development in response to environmental changes, yet the underlying mechanisms are largely unknown. Here we report the identification of two rice leucine-rich repeat receptor-like kinases, Thermo-Sensitive Genic Male Sterile 10 (TMS10) and its close homolog TMS10-Like (TMS10L), which redundantly function in the maintenance of the tapetal cell layer and microspore/pollen viability under normal temperature conditions with TMS10 playing an essential role in higher temperatures (namely, 28 °C). tms10 displays male sterility under high temperatures but male fertility under low temperatures, and the tms10 tms10l double mutant shows complete male sterility under both high and low temperatures. Biochemical and genetic assays indicate that the kinase activity conferred by the intracellular domain of TMS10 is essential for tapetal degeneration and male fertility under high temperatures. Furthermore, indica or japonica rice varieties that contain mutations in TMS10, created by genetic crosses or genome editing, also exhibit thermo-sensitive genic male sterility. These findings demonstrate that TMS10 and TMS10L act as a key switch in postmeiotic tapetal development and pollen development by buffering environmental temperature changes, providing insights into the molecular mechanisms by which plants develop phenotypic plasticity via genotype-environment temperature interaction. TMS10 may be used as a genetic resource for the development of hybrid seed production systems in crops.

AN OVERVIEW OF POLLEN AND ANTHER WALL DEVELOPMENT IN Catalpa bignonioides WALTER (BIGNONIACEAE)

Anther development in Catalpa bignonioides Walter was investigated from the sporogenous cell to the mature pollen grain stages to determine whether the pollen and anther wall development follows the basic scheme in angiosperms. In order to follow pollen ontogeny through successive stages of pollen development, anthers at different developmental stages were embedded in epon according to the usual method, and semi-thin sections, taken from the epon embedded anthers, were stained with toluidine blue for general histological observations under light microscopy. The young anther wall of C. bignonioides consists of four layers; from the exterior, the epidermis, endothecium, middle layer, and a secretory tapetum. The tapetum is dual in origin and dimorphic. Ubisch bodies were observed on the inner tangential walls of the tapetal cells. The number of the anther wall layers changes depending on the developmental stage and region of the anther. In contrast to the other anther wall layers , epidermis and endothecium layers remain intact until anthesis. Endothecial cells enlarge and develop thickenings at maturity. During microspore development , meiocytes undergo meiosis and simultaneous cytokinesis leading to the formation of permanent tetrahedral, isobilateral and rarely linear tetrads. Pollen tetrads are shed from the anther as compound pollen grains. Results of the study revealed that pollen and anther wall development in C. bignonioides follows the basic scheme in angiosperms.

Acetolactate Synthase-Inhibiting Gametocide Amidosulfuron Causes Chloroplast Destruction, Tissue Autophagy, and Elevation of Ethylene Release in Rapeseed.

Background: Acetolactate synthase (ALS)-inhibiting herbicides amidosulfuron (Hoestar) is an efficient gametocide that can induce male sterility in rapeseed (Brassica napus L.). We conducted an integrated study of cytological, transcriptomic, and physiological analysis to decipher the gametocidal effect of amidosulfuron.Results: In the first several days after exposure to amidosulfuron at a gametocidal dose of ca. 1 μg per plant, the plants showed the earliest symptoms including short retard of raceme elongation, slight chlorosis on leaf, and decrease of photosynthesis rate. Chloroplasts in leaf and anther epidermis, and tapetal plastids were deformed. Both tapetal cell and uni-nucleate microspore showed autophagic vacuoles and degenerated quickly. The amidosulfuron treatment caused reduction of photosynthetic rate and the contents of leaf chlorophyll, soluble sugar and pyruvate, as well as content alteration of several free amino acids in the treated plants. A comparison of transcriptomic profiling data of the young flower buds of the treated plants with the control identified 142 up-regulated and 201 down-regulated differential expression transcripts with functional annotations. Down-regulation of several interesting genes encoding PAIR1, SDS, PPD2, HFM1, CSTF77, A6, ALA6, UGE1, FLA20, A9, bHLH91, and putative cell wall protein LOC106368794, and up-regulation of autophagy-related protein ATG8A indicated functional abnormalities about cell cycle, cell wall formation, chloroplast structure, and tissue autophagy. Ethylene-responsive transcription factor RAP2-11-like was up-regulated in the flower buds and ethylene release rate was also elevated. The transcriptional regulation in the amidosulfuron-treated plants was in line with the cytological and physiological changes.Conclusions: The results suggested that metabolic decrease related to photosynthesis and energy supply are associated with male sterility induced by amidosulfuron. The results provide insights into the molecular mechanisms of gametocide-induced male sterility and expand the knowledge on the transcriptomic complexity of the plants exposure to sulfonylurea herbicide.

Ontogeny of permanent tetrads in Gardenia jasminoides (Rubiaceae) provides insight into pollen evolution

Tetrad pollen ontogeny of Gardenia jasminoides was studied to explore potential insights for the ontogeny of compound pollen and the evolution of pollen in Rubiaceae. Its developmental changes were examined by light microscope, scanning and transmission electron microscope. After meiosis, the cleavage wall is centrifugally formed, which is a first report in Rubiaceae. After tetrads releasing, intine begins to accumulate at non-apertural regions, and then extends to apertural regions. This formation pattern is underreported in angiosperms. The mature pollen grains are united in permanent tetrahedral tetrads, and they are bound by simple cohesion, with complete intermeshing of tectum and columella. Detailed development process of the intermeshing is described for the first time in Rubiaceae. Apertures are pairly arranged as Fischer's rule, and protruding onci are formed at apertural regions. The tapetum is of secretory type and an occasional movement of tapetal cells is found at uninucleate microspore stage. This study has revealed valuable information for understanding the great diversity of pollen characters in Rubiaceae.

Cytological and Transcriptomic Analyses Reveal Important Roles of CLE19 in Pollen Exine Formation.

The CLAVATA3/ESR-RELATED (CLE) peptide signals are required for cell-cell communication in several plant growth and developmental processes. However, little is known regarding the possible functions of the CLEs in the anther. Here, we show that a T-DNA insertional mutant, and dominant-negative (DN) and overexpression (OX) transgenic plants of the CLE19 gene, exhibited significantly reduced anther size and pollen grain number and abnormal pollen wall formation in Arabidopsis (Arabidopsis thaliana). Interestingly, the DN-CLE19 pollen grains showed a more extensively covered surface, but CLE19-OX pollen exine exhibited clearly missing connections in the network and lacked separation between areas that normally form the lacunae. With a combination of cell biological, genetic, and transcriptomic analyses on cle19, DN-CLE19, and CLE19-OX plants, we demonstrated that CLE19-OX plants produced highly vacuolated and swollen aborted microspores (ams)-like tapetal cells, lacked lipidic tapetosomes and elaioplasts, and had abnormal pollen primexine without obvious accumulation of sporopollenin precursors. Moreover, CLE19 is important for the normal expression of more than 1,000 genes, including the transcription factor gene AMS, 280 AMS-downstream genes, and other genes involved in pollen coat and pollen exine formation, lipid metabolism, pollen germination, and hormone metabolism. In addition, the DN-CLE19(+/+) ams(-/-) plants exhibited the ams anther phenotype and ams(+/-) partially suppressed the DN-CLE19 transgene-induced pollen exine defects. These findings demonstrate that the proper amount of CLE19 signal is essential for the normal expression of AMS and its downstream gene networks in the regulation of anther development and pollen exine formation.

Gene Regulatory Network for Tapetum Development in Arabidopsis thaliana.

In flowering plants, male gametophyte development occurs in the anther. Tapetum, the innermost of the four anther somatic layers, surrounds the developing reproductive cells to provide materials for pollen development. A genetic pathway of DYT1-TDF1-AMS-MS188 in regulating tapetum development has been proven. Here we used laser microdissection and pressure catapulting to capture and analyze the transcriptome data for the Arabidopsis tapetum at two stages. With a comprehensive analysis by the microarray data of dyt1, tdf1, ams, and ms188 mutants, we identified possible downstream genes for each transcription factor. These transcription factors regulate many biological processes in addition to activating the expression of the other transcription factor. Briefly, DYT1 may also regulate early tapetum development via E3 ubiquitin ligases and many other transcription factors. TDF1 is likely involved in redox and cell degradation. AMS probably regulates lipid transfer proteins, which are involved in pollen wall formation, and other E3 ubiquitin ligases, functioning in degradating proteins produced in previous processes. MS188 is responsible for most cell wall-related genes, functioning both in tapetum cell wall degradation and pollen wall formation. These results propose a more complex gene regulatory network for tapetum development and function.

An atypical strictosidine synthase, OsSTRL2, plays key roles in anther development and pollen wall formation in rice

Strictosidine synthase (STR) plays an important role in the biosynthesis of terpenoid indole alkaloids (TIAs) and is expressed in a range of active meristematic tissues of higher plants. STR proteins are involved in different physiological and biochemical pathways. However, the function of STR proteins in rice development remains poorly understood. In this study, we identified 21 possible STR-like (OsSTRL) family members in rice genome and found that only one gene, OsSTRL2, exhibited a pre-emergency specific florescence expression pattern. Tissue-specific expression profile analysis, β-glucuronidase histochemical (GUS) staining and RNA in situ hybridization confirmed that OsSTRL2 was highly expressed in tapetal cells and microspores. Comparative protein sequence analysis indicated that OsSTRL2 lacked the key catalytic residue found in a typical STR (STR1), although it possessed conserved β-propellers and α-helices formed the basic structure of STR1. OsSTRL2 knockout mutant resulted to male sterility because of the defects in anther development and pollen wall formation. Subcellular localization of OsSTRL2-YFP revealed that the OsSTRL2 protein was primarily localized in the endoplasmic reticulum (ER). Therefore, OsSTRL2 is an atypical strictosidine synthase that plays crucial roles in regulating anther development and pollen wall formation in rice.

Carbonic Anhydrases Function in Anther Cell Differentiation Downstream of the Receptor-Like Kinase EMS1.

Plants extensively employ leucine-rich repeat receptor-like kinases (LRR-RLKs), the largest family of RLKs, to control a wide range of growth and developmental processes as well as defense responses. To date, only a few direct downstream effectors for LRR-RLKs have been identified. We previously showed that the LRR-RLK EMS1 (EXCESS MICROSPOROCYTES1) and its ligand TPD1 (TAPETUM DETERMINANT1) are required for the differentiation of somatic tapetal cells and reproductive microsporocytes during early anther development in Arabidopsis thaliana Here, we report the identification of β-carbonic anhydrases (βCAs) as the direct downstream targets of EMS1. EMS1 biochemically interacts with βCA proteins. Loss of function of βCA genes caused defective tapetal cell differentiation, while overexpression of βCA1 led to the formation of extra tapetal cells. EMS1 phosphorylates βCA1 at four sites, resulting in increased βCA1 activity. Furthermore, phosphorylation-blocking mutations impaired the function of βCA1 in tapetal cell differentiation; however, a phosphorylation mimic mutation promoted the formation of tapetal cells. βCAs are also involved in pH regulation in tapetal cells. Our findings highlight the role of βCA in controlling cell differentiation and provide insights into the posttranslational modification of carbonic anhydrases via receptor-like kinase-mediated phosphorylation.

Polysaccharides and lipids in the developing anthers of Hong Kong orchid tree (Bauhinia blakeana)

Key message Microspore ofBauhinia blakeanabegins to accumulate lipids, and starches appear following bicellular pollen development. The development of tapetal cells ofB. blakeanais unusual in different stage.

Rice fatty acyl-CoA synthetase OsACOS12 is required for tapetum programmed cell death and male fertility.

Loss of function mutation of rice OsACOS12 impairs lipid metabolism-mediated anther cuticle and pollen wall formation, and interferes with tapetum programmed cell death, leading to male sterility. Acyl-CoA Synthetase (ACOS) is one of the enzymes activating fatty acids for various metabolic functions in plants. Here, we show that OsACOS12, an orthologue of Arabidopsis ACOS5 in rice, is crucial for rice fertility. Similar to acos5, osaocs12 mutant had no mature pollen. But unlike acos5, osaocs12 produced defective anthers lacking cutin and Ubisch bodies on the epidermal and inner surfaces, respectively, and delayed programmed cell death (PCD)-induced tapetum degradation. Those phenotypic changes were evident at stage 10, during which OsACOS12 had its maximum expression in tapetal cells and microspores. Chemical analysis revealed that the levels of anther cuticular lipid components (wax and cutin monomers) were significantly reduced in osaocs12, while the expression levels of three known lipid biosynthetic genes were unchanged. Recombinant OsACOS12 enzyme was shown to catalyze the conversion of C18:1 fatty acid to C18:1 CoA in vitro. Phylogenetic analysis indicated that OsACOS12 is an ancient and conserved enzyme associated with the plant's colonization to earth. Collectively, our study suggests that OsACOS12 is an ancient enzyme participating in a conserved metabolic pathway for diversified biochemical functions to secure male reproduction in plants.