Plant and Cell Physiology, 1997, Vol. 38, No. 3 304-311
© 1997
Identification of Stylar RNases Associated with Gametophytic Self-Incompatibility in Almond (Prunus dulcis)
1 Faculty of Agriculture, Kyoto University Kyoto, 606-01 Japan
2 Faculty of Horticulture, Chiba University Chiba, 271 Japan
3 School of Agricultural Sciences, Nagoya University Nagoya, 464-01 Japan
4 Department of Pomology, University of California Davis, CA 95616, U.S.A.
Correspondence to Ryutaro Tao (FAX: 075-753-6068, E-mail: rtao{at}kais.kyoto-u.ac.jp).
Stylar proteins of 13 almond (Prunus dulcis) cultivars with known S-genotypes were surveyed by IEF and 2D-PAGE combined with immunoblot and N-terminal amino acid sequence analyses to identify S-RNases associated with gametophytic self-incompatibility (SI) in this plant species. RNase activities corresponding to Sa and Sb, two of the four S-alleles tested, were identified by IEF and RNase activity staining. The Sa-RNase band reacted with the anti-S4serum prepared from Japanese pear (Pyrus serotina); no reaction with the antiserum was observed with the sbRNase band. When the sa-RNase band was excised from an IEF gel stained for RNase activity, subjected to SDS-PAGE, and detected by immunoblotting, it appeared that this band consisted of a single protein that reacted with the anti-s4serum with Mr of about 28 kDa. With 2D-PAGE and silver staining of the stylar extracts, all four S-proteins could be successfully distinguished from each other in the highly basic zone of the gel. Although Sb-, Sc-, and Sdproteins had roughly the same Mr of about 30 kDa, the Sc-protein seemed to be slightly smaller than the Sb-protein and slightly larger than the Saprotein. In 2D-PAGE profiles as well, the Sa-protein had Mr of about 28 kDa, apparently smaller than the other three proteins. A bud sport, in which one of the two S-alleles of the original cultivar is impaired, was visualized as a loss of Scprotein, which is consistent with the previous pollination study. All four S-proteins reacted with the anti-S4serum, probably because of the differing conformations of these S-proteins in the IEF and 2D-PAGE gels. The Sa-protein in 2D-PAGE appeared to be identical to Sa-RNase in IEF; both bad the same Mr and were reactive with the anti-S4-serum. N-terminal amino acid sequence analysis of the four 5-proteins revealed that they were highly homologous to each other and similar to the 5-RNases of Malus, Pyrus, Scrophulariaceae, and Solanaceae. Taken together, RNases in the style are strongly suggested to be associated with the gametophytic SI of al- mond. This is the first report identofiying and characterizing S-RNase in almond.
(Received July 11, 1996; Accepted December 26, 1996)
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