Identification and partial amino acid sequences of seven S-RNases associated with self-incompatibility of Japanese pear, Pyrus pyrifolia Nakai

Abstract

S-allele-specific proteins (S-proteins) were separated and identified by two-dimensional (2D) gel electrophoresis from the style extract of 14 cultivars of Japanese pear, Pyrus pyrifolia Nakai, which exhibits gametophytic self-incompatibility. These S-proteins were 30-32 kDa basic proteins with putative pIs of 9.6-10.1 and were distinct from the other proteins, which were common for all cultivars examined. Each S-protein was assigned to a given S-genotype based on electrophoretic mobility and the partial amino acid sequence. For S1- to S7-proteins, five different N-terminal amino acid sequences sharing the YFQFTQQY sequence were determined. Since the same N-terminal amino acid sequences were found for both S1- and S7-proteins, and for S3- and S5-proteins, the two S-proteins of each pair were distinguished based on their electrophoretic behavior. The internal amino acid sequences of S2- and S4-proteins, determined for Achromobacter protease I (API) digests, revealed that these proteins are S2- and S4-RNases, respectively. In the cultivar Nijisseiki, these two RNases were expressed from the white bud to mature flower stages when the cultivar acquires and enforces self-incompatibility. Osa-nijisseiki, a self-compatible mutant of Nijisseiki, produced S2-RNase, but did not produce S4-RNase. The absence of S4-RNase was also observed in self-compatible offsprings derived from Osa-Nijisseiki. These results suggest that Japanese pear in the family Rosaceae possesses a gametophytic self-incompatibility system involving an S-RNase, and that a reduction or lack of expression of S4-RNase in the style is responsible for the self-compatibility of Osa-Nijisseiki.