Molecular cloning and nucleotide sequences of cDNAs encoding S-allele specific stylar RNases in a self-incompatible cultivar and its self-compatible mutant of Japanese pear, Pyrus pyrifolia Nakai

Abstract

The genes encoding three RNases were cloned from the style of a self-incompatible cultivar, Nijisseiki (S2S4), and its self-compatible mutant, Osa-Nijisseiki (S2S4sm, sm means stylar part mutant), of Japanese pear. For Nijisseiki, cDNAs coding for two S-RNase (S2-RNase and S4-RNase) and an RNase unrelated to self-incompatibility (non-S-RNase) were cloned from the stylar cDNA library. The cDNAs coding for S2-RNase, S4-RNase, and non-S-RNase include 678-, 684-, and 681-bp open reading frames, respectively. Their deduced amino acid sequences were composed of signal peptides and mature RNases (201-203 residues) which were verified by partial amino acid sequencing. The primary structures of mature proteins revealed that these RNases are of the RNase T2 type; only the two S-RNases have several potential N-glycosylation sites and 60% of their amino acid residues are identical, compared with 25% sequence identity with the non-S-RNase. Such a distinct difference in the primary structures between S-RNases and non-S-RNase has not previously been reported and may be a feature typical of S-RNases in the family Rosaceae. Similar experiments were performed for Osa-Nijisseiki. The cDNAs coding for S2-RNase and non-S-RNase were similarly cloned from the stylar cDNA library. However, the cDNA coding for S4-RNase was neither amplified by PCR nor cloned from the library, suggesting that the mutation of self-incompatible Nijisseiki to self-compatible Osa-Nijisseiki is due to a failure of expression of S4-RNase. These results lead to the idea that Osa-Nijiisseiki is a variant of Nijisseiki in which the S4-allelic gene in the S-locus is exclusively mutated or deleted, causing severely impaired or suppressed expression of its gene product, S4-RNase, at the style.