OsZNHIT3 functions as a transcriptional activator to regulate tapetal programmed cell death and exine formation in...

Proper timing of programmed cell death (PCD) in the tapetum and the deposition of pollen wall are crucial for gametophyte development. Although several genes involved in these processes have been identified, the regulatory mechanisms remain poorly understood. We isolated and characterized a rice male-sterile mutant, exine abnormal 1 (exa1), which exhibits pale anthers and aborted pollen. The exa1 mutant displays delayed tapetal PCD and defective pollen wall patterning, including thinner sexine and thicker nexine. EXA1 (Exine Abnormal 1), preferentially expressed in the tapetum and microspores, encodes a novel Zf-HIT (Zinc finger-High Temperature) family protein, OsZNHIT3. Nucleus and cytoplasm dual-localized EXA1 functions as a transcriptional activator via its previously undefined C-terminal domain. Phenotypic similarities in rice expressing EXA1-SRDX (SUPERMAN Repressive Domain X) fusion proteins further support its role as a transcriptional activator. PTC1 (Persistent Tapetal Cell 1), EAT1 (Eternal Tapetum 1), and their downstream genes were significantly downregulated in exa1 mutant anthers. Introducing pEXA1:PTC1 or pEXA1:EAT1 into the exa1 mutant partially rescued anther appearance, pollen morphology, and sexine-nexine ratio. This suggests that EXA1 acts genetically upstream of PTC1 and EAT1 in the regulatory pathway. Our study reveals a novel and essential role for EXA1 as a transcriptional activator in rice anther development.