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The plant pollen wall protects the male gametophyte from various biotic and abiotic stresses. The formation of a unique pollen wall structure and elaborate exine pattern is a well-organized process, which needs coordination between reproductive cells and the neighboring somatic cells. However, molecular mechanisms underlying this process remain largely unknown. Here, we report a rice male-sterile mutant (l94) that exhibits defective pollen exine patterning and abnormal tapetal cell development. MutMap and knockout analyses demonstrated that the causal gene encodes a type-G non-specific lipid transfer protein (OsLTPL94). Histological and cellular analyses established that OsLTPL94 is strongly expressed in the developing microspores and tapetal cells, and its protein is secreted to the plasma membrane. The l94 mutation impeded the secretory ability of OsLTPL94 protein. Further in vivo and in vitro investigations supported the hypothesis that ETERNAL TAPETUM 1 (EAT1), a basic helix-loop-helix transcription factor (bHLH TF), activated OsLTPL94 expression through direct binding to the E-box motif of the OsLTPL94 promoter, which was supported by the positive correlation between the expression of EAT1 and OsLTPL94 in two independent eat1 mutants. Our findings suggest that the secretory OsLTPL94 plays a key role in the coordinated development of tapetum and microspores with the regulation of EAT1., (© 2021 Society for Experimental Biology and John Wiley & Sons Ltd.)

Starch accumulation is key for the maturity of rice pollen grains; however, the regulatory mechanism underlying this process remains unknown. Here, we have isolated a male-sterile rice mutant, abnormal pollen 1 ( ap1 ), which produces nonviable pollen grains with defective starch accumulation. Functional analysis revealed that AP1 encodes an active L-type lectin receptor-like kinase (L-LecRLK). AP1 is localized to the plasma membrane and its transcript is highly accumulated in pollen during the starch synthesis phase. RNA-seq and phosphoproteomic analysis revealed that the expression/phosphorylation levels of numerous genes/proteins involved in starch and sucrose metabolism pathway were significantly altered in the mutant pollen, including a known rice UDP-glucose pyrophosphorylase (OsUGP2). We further found that AP1 physically interacts with OsUGP2 to elevate its enzymatic activity, likely through targeted phosphorylation. These findings revealed a novel role of L-LecRLK in controlling pollen maturity via modulating sucrose and starch metabolism., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

Meiosis is critical for sexual reproduction and the generation of new allelic variations in most eukaryotes. In this study, we report the isolation of a meiotic gene, DLC1, using a map-based cloning strategy. The dlc1 mutant is sterile in both male and female gametophytes due to an earlier defect in the leptotene chromosome and subsequent abnormalities at later stages. DLC1 is strongly expressed in the pollen mother cells (PMCs) and tapetum and encodes a nucleus-located rice type-B response regulator (RR) with transcriptional activity. Further investigations showed that DLC1 interacts with all five putative rice histidine phosphotransfer proteins (HPs) in yeast and planta cells, suggesting a possible participation of the two-component signalling systems (TCS) in rice meiosis. Our results demonstrated that DLC1 is required for rice meiosis and fertility, providing useful information for the role of TCS in rice meiosis., (© 2019 The Authors The Plant Journal © 2019 John Wiley & Sons Ltd.)

Key Message: OsPKS2, the rice orthologous gene 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.

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.

SUMMARY: Pollen development includes a series of biological events that require precise gene regulation. Although several transcription factors (TFs) have been shown to play roles in maintaining pollen fertility, the major regulatory networks underlying tapetum development and pollen wall formation are largely unknown. Herein, we report that ABERRANT MICROSPORE DEVELOPMENT1 (AMD1), a protein annotated previously as unknown protein, is required for tapetum development and pollen exine patterning in rice (Oryza sativa L.). AMD1 encodes a grass‐specific protein exhibiting transactivation activity in the nucleus and is spatiotemporally expressed in the tapetum and microspores during pollen development. Further biochemical assays indicate that AMD1 directly activates the transcription of DEFECTIVE POLLEN WALL (DPW) and POLYKETIDE SYNTHASE2 (OsPKS2), which are both implicated in sporopollenin biosynthesis during exine formation. Additionally, AMD1 directly interacts with TAPETUM DEGENERATION RETARDATION (TDR), a key TF involved in the regulation of tapetum degradation and exine formation. Taken together, we demonstrate that AMD1 is an important regulatory component involved in the TDR‐mediated regulatory pathway to regulate sporopollenin biosynthesis, tapetum degradation, and exine formation for pollen development. Our work provides insights into the regulatory network of rice sexual reproduction and a useful target for genetic engineering of new male‐sterile lines for hybrid rice breeding. [ABSTRACT FROM AUTHOR]

Arabinogalactan proteins (AGPs) are widely distributed in plant cells. Fasciclin‐like AGPs (FLAs) belong to a subclass of AGPs that play important roles in plant growth and development. However, little is known about the biological functions of rice FLA. Herein, we report the identification of a male‐sterile mutant of DEFECTIVE EXINE AND APERTURE PATTERNING1 (DEAP1) in rice. The deap1 mutant anthers produced aberrant pollen grains with defective exine formation and a flattened aperture annulus and exhibited slightly delayed tapetum degradation. DEAP1 encodes a plasma membrane‐associated member of group III plant FLAs and is specifically and temporally expressed in reproductive cells and the tapetum layer during male development. Gene expression studies revealed reduced transcript accumulation of genes related to exine formation, aperture patterning, and tapetum development in deap1 mutants. Moreover, DEAP1 may interact with two rice D6 PROTEIN KINASE‐LIKE3s (OsD6PKL3s), homologs of a known Arabidopsis aperture protein, to affect rice pollen aperture development. Our findings suggested that DEAP1 is involved in male reproductive development and may affect exine formation and aperture patterning, thereby providing new insights into the molecular functions of plant FLAs in male fertility. [ABSTRACT FROM AUTHOR]

Pollen development in flowering plants is critical for male reproductive success. The pollen wall that protects the pollen from various environment stresses and bacterial infections plays an essential role in pollen development. The formation of pollen wall is associated with the biosynthesis and transport of sporopollenin components. ACOS5 in Arabidopsis encodes an acyl-CoA synthetase 5 required for sporopollenin biosynthesis. We identified the rice homolog of ACOS5 as OsACOS12. The CRISPR/Cas9-mediated OsACOS12 knockout mutant has complete male sterility due to a defect in pollen wall formation. β-Glucuronidase reporter gene analysis and RNA in situ hybridization indicated that OsACOS12 was specifically expressed in tapetum and microspores. The subcellular localization of OsACOS12-YFP demonstrated that OsACOS12 protein was primarily localized in the endoplasmic reticulum and nucleus. Our results suggest that OsACOS12 plays a critical and conserved role in pollen wall formation and pollen development and has implications in rice breeding. [ABSTRACT FROM AUTHOR]

Pollen development plays crucial roles in the life cycle of higher plants. Here we characterized a rice mutant with complete male-sterile phenotype, pollen-less 1 (pl1). pl1 exhibited smaller anthers with arrested pollen development, absent Ubisch bodies, necrosis-like tapetal hypertrophy, and smooth anther cuticular surface. Molecular mapping revealed a synonymous mutation in the fourth exon of PL1 co-segregated with the mutant phenotype. This mutation disrupts the exon-intron splice junction in PL1, generating aberrant mRNA species and truncated proteins. PL1 is highly expressed in the tapetal cells of developing anther, and its protein is co-localized with plasma membrane (PM) and endoplasmic reticulum (ER) signal. PL1 encodes an integrin-α FG-GAP repeat-containing protein, which has seven β-sheets and putative Ca2+-binding motifs and is broadly conserved in terrestrial plants. Our findings therefore provide insights into both the role of integrin-α FG-GAP repeat-containing protein in rice male fertility and the influence of exonic mutation on intronic splice donor site selection. [ABSTRACT FROM AUTHOR]