Characteristics Of S-RNase And Its Mechanisms Related To Growing Inhibition Of Self-Pollen Tube In Japanese Apricot (Prunus Mume)

Self-incompatibility （SI） is a genetic mechanism employed by flowering plants toprovent inbreeding and promoting out-crossing, which involves a complex set of cell-cellinteractions between the pistil and the pollen, thence, a model system for studying ofintercellular information transmission, celll-cell recognition and gene spatial-temporalexpression. Extensive studies have been carried out in species of Solanaceae, Rosaceae andScrophulariaceae, the majority of which displays an S-RNase-mediated gametophyticself-incompatibility （GSI）. Information available indicates that GSI response is under thegenetic control of a single multi-allelic locus, the S-locus, which contains at least twoseparate genes, S-RNase and SFB/SLF, Controlling female and male specificity respectively,hence the name ’S-haplotype’ to describe variants of the S-locus. Recognition betweenS-RNase and SFB/SLF of the same S-haplotype triggers an SI reaction.Japanese apricot （Prunus mume） is a commercially important fruit crop in china, whereasexhibits S-RNase-based gametophytic self-incompatibility, and most cultivars requirepollinators inter-planted to ensure adequate pollination.Japanese apricot ’Xiyeqing’, ’Oushuku’ and the others were used as materials. Thecharacteristics of S-RNase and the mechanisms of inhibitting pollen tube growth byS-RNase were investigated with modem biological research method. The main results weresummarized as follows:1. Self-incompatibility strength of Japanese apricot 10 cultivarsThrough self-pollinated fruit setting rate, and numbers of pollen tube in base style, andS-RNase content, the varieties of self-incompatibility strength was determined. As strong:’Nankou’; middle: ’Oushuku’, ’Gessekai’, ’Xiyeqing’ and ’Nanhong’; Weak: ’Bungo’,’Ruantiaohongmei’ and ’Xiaoyezhugan’. ’Tougorou’ and ’Hachirou’ is self-compatibility （SC）,but the expression of S-RNase is normal.2. Self-incompatibility strength of floral different developmental stages in Japanese apricot S-RNase began to synthesize from - 6 days after anthesis （DAA）, and the largestsynthesis at -2～-3 DAA, then slowly degraded after anthesis in Japanese apricot cultivar’Xiyeqing’. Fruit set percentage after self-pollination and self-tube numbers in the base ofstyle shows the same results: the SI strength at -4 DAA is weak, as well as thedevelopment of a gradual increase, the strongest at anthesis, then weak after anthesis. Theeffective period of self-pollination was -2 DAA.S-RNase content showed negative relations to fruit set (r=0.9640^**), and the number ofpollen tubes arriving at style base (r=0.9380^**) respectively. The S-RNase content is themain reason for SI strength of Janaese apricot was suggested.3. Content and purification of S-RNase in Japanese apricotThe S-RNase of cultivar ’Xiyeqing’ specific expresses in the pistil, which is abundantlyin style and a few in ovary, but failed to detect expression in anther, petal, sepal, receptacle,bud and young fruit. Application of CM and SP ion chromatography column can be purifiedthe S-RNase of Japanese apricot, 86ï¼…-96ï¼…purity. The average volume of extraction is130.9μg from lg of styles. The S-RNase molecular weight is 32kD around.4. Characteristics of pollen germination and tube growth of Japanese apricot in vitroThe optimum liquid medium for pollen germination and pollen tube growth of’Xiyeqing’ was 30mmol/L MES （pH6.5） buffer containing 20ï¼…sucros, 0.01ï¼…H₃BO₃,20ï¼…PEG-4000, 0.03ï¼…Ca（NO₃）2·4H₂O, 0.02ï¼…MgSO₄·7H₂O. Rate of germination was45.0ï¼…and length of pollen tube was 597.2μm. In addition, the suitable for culture pollen:the density and temperature of 20-80 grain perμl, 25℃.5. S-RNase inhibited the pollen germination and pollen tube growth in vitroThe S-RNase of ’Gessekai’ at the activity of 0.0028 U·μl^-1 can inhibit the self-pollengermination and tube growth specificly, but such specific inhibition disappeared at 35℃. Ifthe activity is greater than 0.0046 U·μl^-1, it started to inhibit cross-pollen germination andtube growth. The exogenous RNase A can inhibit pollen germination and pollen tubegrowth, but stylar crude proteins without S-RNase does not.6. The degradation of tube RNA by S-RNase in vitroThe RNA of self-pollen tube was degraded by S-RNase in vitro in Japanese apricotspecificly. But as time has been extended the RNA of cross-pollen tube was degraded too.The substrate for S-RNases may not specificity, but the degradation rate may be specificitywas suggested. With the rising of temperature or increase of the S-RNase activity thisspecific degradation drops or even disappears. The RNA of pollen tube can be degraded by stylar crude proteins without S-RNase and exogenous RNase A without specificity.Therefore, specific degradation of self-tube RNA caused self-pollen tube growth slowerthan cross-pollen tube was proposed.In this paper, the self-incompatibility strength in different Japanese apricot cultivar andfloral stage was first analysed. From the purification and characteristic and function ofS-RNase to systematic investigate self-incompatibility in Japanese apricot, and thepurification of S-RNase from Japanese apricot was first studied. The results furtherrevealed that Japanese apricot is gametophytic self-incompatibility （GSI） based on S-RNase.The SI mechanism is specific degradation of pollen tube RNA by stylar S-RNase causedpollen tube growth rest was proposed, which further enriching the mechanism of GSI in theRosaceae.