Premature senescence of leaves causes a reduced yield and quality of rice by affecting plant growth and development. The regulatory mechanisms underlying early leaf senescence are still unclear. The Leaf senescence 1 (LS1) gene encodes a C2H2-type zinc finger protein that is localized to both the nucleus and cytoplasm. In this study, we constructed a rice mutant named leaf senescence 1 (ls1) with a premature leaf senescence phenotype using CRISPR/Cas9-mediated editing of the LS1 gene. The ls1 mutants exhibited premature leaf senescence and reduced chlorophyll content. The expression levels of LS1 were higher in mature or senescent leaves than that in young leaves. The contents of reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD) were significantly increased and catalase (CAT) activity was remarkably reduced in the ls1 plants. Furthermore, a faster decrease in pigment content was detected in mutants than that in WT upon induction of complete darkness. TUNEL and staining experiments indicated severe DNA degradation and programmed cell death in the ls1 mutants, which suggested that excessive ROS may lead to leaf senescence and cell death in ls1 plants. Additionally, an RT-qPCR analysis revealed that most senescence-associated and ROS-scavenging genes were upregulated in the ls1 mutants compared with the WT. Collectively, our findings revealed that LS1 might regulate leaf development and function, and that disruption of LS1 function promotes ROS accumulation and accelerates leaf senescence and cell death in rice.

Plant proline-rich proteins (PRPs) are cell wall proteins that are widely distributed in plants. Previous studies have shown that these proteins play a crucial role in adversity stress processes, but their function in the regulation of pollen fertility in rice remains unknown. In this study, we identified that OsPRP1 contains a Pollen_Ole-e_I allergenic structural domain, obtained the OsPRP1 quadruple mutant named osprp1.1/1.2/1.3/1.4, and observed significant reductions in pollen fertility, seed-setting rates, and the deformation and collapse of microspores during the late stages of pollen development. RNA-Seq analysis indicated the down-regulation of genes involved in anther development in osprp1.1/1.2/1.3/1.4, suggesting that OsPRP1 plays a role in regulating pollen fertility. In conclusion, a loss of function in OsPRP1.1/1.2/1.3/1.4 leads to decreased pollen fertility and seeding rates, which not only expands the functional spectrum of plant PRP genes but also provides new theoretical insight into the mechanism of fertility regulation in rice.