Five Family Members Are Identified as Senescence-Related Genes in Rice by Reverse Genetics Approach.

A total of 16 genes of were identified in our previous work, but their functions remain unclear. In this study, 13 members were knocked out using the CRISPR/cas9 gene-editing technology. After screening phenotype characterization of CRISPR/Cas9 mutants compared to WT, five mutants exhibited a stay-green phenotype at 30 days after heading. Moreover, increased grain size and grain weight occurred in the , and lines, while declined grain weight appeared in the and mutants. The transcript levels of several senescence-associated genes (SAGs), chlorophyll degradation-related genes (CDGs), as well as WRKY members were differentially decreased in the five stay-green mutants compared to WT. Five mutants also exhibited a stay-green phenotype when the detached leaves were incubated under darkness for 4 days. and were significantly upregulated, while and were significantly downregulated in both and compared to WT. Furthermore, these five displayed strong transcriptional activation activity and were located in the nucleus. Most of the genes were downregulated in the , and mutants, but upregulated in the and mutants, indicating coordinated regulation among members. These results suggest that , and are senescence-associated genes, involved in the senescence and carbon allocation network by modulating other members, member genes, and senescence-related gene expression.