Structural variations of a new fertility restorer gene, Rf20, underlie the restoration of wild abortive-type cytoplasmic male sterility in rice.

The discovery of a wild abortive-type (WA) cytoplasmic male sterile (CMS) line and breeding its restorer line have led to the commercialization of three-line hybrid rice, contributing considerably to global food security. However, the molecular mechanisms underlying fertility abortion and the restoration of CMS-WA lines remain largely elusive. In this study, we cloned a restorer gene, Rf20, following a genome-wide association study analysis of the core parent lines of three-line hybrid rice. We found that Rf20 was present in all core parental lines, but different haplotypes and structural variants of its gene resulted in differences in Rf20 expression levels between sterile and restored lines. Rf20 could restore pollen fertility in the CMS-WA line and was found to be responsible for fertility restoration in some CMS lines under high temperatures. In addition, we found that Rf20 encodes a pentatricopeptide repeat protein that competes with WA352 for binding with COX11. This interaction enhances COX11's function as a scavenger of reactive oxygen species, which in turn restores pollen fertility. Collectively, our study suggests a new action mode for pentatricopeptide repeat proteins in the fertility restoration of CMS lines, providing an essential theoretical basis for breeding robust restorer lines and for overcoming high temperature-induced fertility recovery of some CMS lines.