Gametophyte development in angiosperms occurs within diploid sporophytic structures and requires coordinated development; e.g., development of the male gametophyte pollen depends on the surrounding sporophytic tissue, the tapetum. The mechanisms underlying this interaction remain poorly characterized. The peptide CLAVATA3/EMBRYO SURROUNDING REGION-RELATED 19 (CLE19) plays a “braking” role in preventing the harmful overexpression of tapetum transcriptional regulators to ensure normal pollen development in Arabidopsis. However, the CLE19 receptor is unknown. Here, we show that CLE19 interacts directly with the PXY-LIKE1 (PXL1) ectodomain and induces PXL1 phosphorylation. PXL1 is also required for the function of CLE19 in maintaining the tapetal transcriptional regulation of pollen exine genes. Additionally, CLE19 induces the interactions of PXL1 with SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE (SERK) coreceptors required for pollen development. We propose that PXL1 and SERKs act as receptor and coreceptor, respectively, of the extracellular CLE19 signal, thereby regulating tapetum gene expression and pollen development.

The pollen wall is a specialized extracellular cell wall that protects male gametophytes from various environmental stresses and facilitates pollination. Here, we reported that bHLH010 and bHLH089 together are required for the development of the pollen wall by regulating their specific downstream transcriptional and metabolic networks. Both the exine and intine structures of bhlh010 bhlh089 pollen grains were severely defective. Further untargeted metabolomic and transcriptomic analyses revealed that the accumulation of pollen wall morphogenesis-related metabolites, including polysaccharides, glyceryl derivatives, and flavonols, were significantly changed, and the expression of such metabolic enzyme-encoding genes and transporter-encoding genes related to pollen wall morphogenesis was downregulated in bhlh010 bhlh089 mutants. Among these downstream target genes, CSLB03 is a novel target with no biological function being reported yet. We found that bHLH010 interacted with the two E-box sequences at the promoter of CSLB03 and directly activated the expression of CSLB03. The cslb03 mutant alleles showed bhlh010 bhlh089–like pollen developmental defects, with most of the pollen grains exhibiting defective pollen wall structures.

The pollen exine serves as a protective barrier and signaling interface essential for male fertility in flowering plants. Its precise patterning depends on coordinated interactions between microspores and tapetal cells. While the CLAVATA3/EMBRYO SURROUNDING REGION‐related 19 (CLE19) peptide has been identified as a microspore‐derived “brake” that restricts tapetal activity to maintain exine developmental homeostasis, how CLE19 integrates with hormonal signaling pathways remains poorly understood. Here, we demonstrate that CLE19 attenuates brassinosteroid (BR) signaling output by engaging a defined BSL–BIN2–BES1 signaling cascade. Through quantitative phosphoproteomic analysis, we identified that CLE19 affects the phosphorylation of multiple BR signaling components, including BSL‐type phosphatases BSL1/2/3, the GSK3‐like kinase BIN2, and the transcription factor BES1. We show that CLE19 is perceived by its receptor PXL1, which directly interacts with BSL‐type phosphatases to activate the GSK3‐like kinase BIN2, leading to phosphorylation of BES1 at serine residues S219 and S223. Functional analyses using phospho‐dead and phospho‐mimic BES1 variants confirm that CLE19‐dependent phosphorylation controls BES1 nuclear export and degradation, ultimately suppressing BR‐responsive transcriptional outputs required for pollen exine patterning. Together, our findings define a peptide–hormone signaling axis that regulates transcription factor activity through post‐translational regulation, providing mechanistic insight into how developmental robustness is maintained via intercellular signal integration in plant reproduction.

Early anther morphogenesis is a crucial process for male fertility in plants, governed by the transcription factor SPL. While the involvement of AGAMOUS (AG) in SPL activation and microsporogenesis initiation is well established, our understanding of the mechanisms governing the spatial distribution and precise expression of SPL during anther cell fate determination remains limited. Here, we present novel findings on the abnormal phenotypes of two previously unreported SPL mutants, spl-4 and spl-5, during anther morphogenesis. Through comprehensive analysis, we identified ARF3 as a key upstream regulator of SPL. Our cytological experiments demonstrated that ARF3 plays a critical role in restricting SPL expression specifically in microsporocytes. Moreover, we revealed that ARF3 directly binds to two specific auxin response elements on the SPL promoter, effectively suppressing AG-mediated activation of SPL. Notably, the arf3 loss-of-function mutant exhibits phenotypic similarities to the SPL overexpression mutant (spl-5), characterized by defective adaxial anther lobes. Transcriptomic analysis revealed differential expression of the genes involved in the morphogenesis pathway in both arf3 and spl mutants, with ARF3 and SPL exhibited opposing regulatory effects on this pathway. Taken together, our study unveils the precise role of ARF3 in restricting the spatial expression and preventing aberrant SPL levels during early anther morphogenesis, thereby ensuring the fidelity of the critical developmental process in plants.

Faithful pollen wall formation requires tight control of both the amplitude and duration of tapetal transcriptional activity. However, how such dynamic restraint is achieved during anther development remains poorly understood. Here, we identify a phosphorylation-dependent regulatory module in which SBI-PP2A-C1/C5 constrains the functional activity window of the tapetal transcription factor bHLH089 to safeguard male fertility. PP2A-C1 and PP2A-C5 physically associate with bHLH089 and limit its transcriptional output by reducing bHLH089 phosphorylation and nuclear retention, thereby restricting its nuclear accumulation and effective activity. Genetic analyses reveal that precise PP2A-C1/C5 dosage is critical for pollen wall integrity, as both reduced PP2A-C function and PP2A-C1/C5 overexpression disrupt tapetal transcriptional homeostasis and compromise male fertility. Mechanistically, Ser28 and Ser58 function as conserved phosphorylation-dependent determinants of bHLH089 nuclear retention and are required for PP2A-C1/C5-sensitive regulation of bHLH089 activity. Together, these findings identify a phosphatase-mediated timing mechanism that integrates transcriptional feedback with post-translational control to constrain bHLH089-driven transcriptional amplification within a self-limiting tapetal activity window required for tapetal homeostasis, pollen wall formation, and male fertility.

Elevated temperatures, including warm ambient temperature (WAT) and high ambient temperature (HAT), profoundly influence plant growth and development. While WAT promotes thermomorphogenesis and stimulates growth, HAT induces stress responses that can inhibit growth. Using Arabidopsis thaliana , we uncovered both shared and distinct molecular mechanisms underlying plant responses to these temperature conditions. We demonstrated that both WAT and HAT activate genes associated with the DNA damage response (DDR), but they differ in timing and intensity. While WAT encourages progression into the S phase of the cell cycle and enhances cell division, HAT induces G2/M arrest, indicating cell cycle inhibition caused by DNA damage accumulation and impaired DDR proteins. Notably, alternative splicing (AS), particularly intron retention (IR), emerged as a pivotal regulatory layer under heat stress. Prolonged high temperatures uniquely enriched IR events in genes involved in DNA repair and chromatin organization. Integrated analyses of AS and transcriptional changes revealed coordinated regulation, with IR events dominating genes that exhibit simultaneous upregulation. These findings highlight AS as an essential regulatory mechanism in plant responses to heat stress. Conserved responses to elevated temperatures in tobacco and tomato further suggest the presence of shared adaptive pathways across plant species. Our results provide novel insights into plant adaptation to fluctuating temperatures and highlight opportunities for engineering heat-resilient crops to mitigate the impacts of climate change.