S-RNase-mediated attenuation of FviPP2Ac-FviRbohH binding promotes ROS accumulation and arrests pollen tube growth in Fragaria viridis

Abstract

Fragaria viridis exhibits S-RNase-based gametophytic self-incompatibility (SI), which is governed by a polymorphic S-locus including two S-RNase alleles along with multiple S-locus F-box (SLF) genes. However, SI reaction in flowers triggers a complex cascade involving numerous genes unlinked to S-locus, the mechanisms underlying their mediation of SI response remain unclear. In this study, a phosphatase FviPP2Ac and a NADPH oxidase FviRbohH were observed to highly express in pollen and relate to reactive oxygen species (ROS) and SI response. Self-pollen tubes exposed to S-RNase exhibited downregulation of FviPP2Ac and upregulation of FviRbohH. FviPP2Ac played a negative role in pollen tube growth, as supplying FviPP2Ac protein with pollen culture resulted in inhibited pollen tube elongation and reduced ROS content at pollen tube tips. Conversely, FviRbohH acted as a positive regulator, since knockdown of FviRbohH expression in pollen using antisense oligodeoxynucleotides (as-ODN) led to growth suppression, accompanied by decreased ROS levels at pollen tube tips. FviPP2Ac bound to S-RNase in an S-haplotype-independent manner, whereas no interaction was detected between FviRbohH and S-RNase. FviPP2Ac interacted with FviRbohH and inhibited its activity, while S-RNase weakened this interaction without S-haplotype selectivity. Furthermore, we proposed a S-RNase-FviPP2Ac-FviRbohH-ROS signaling pathway model, which provides new clue for revealing a regulatory mechanism about ROS production in SI response of self-pollination in Fragaria viridis.