Identification Of The SCF Complex In The Pear Self-Incompatibility

SI is a universal mechanism in angiosperms that prevents inbreeding and promotes biological diversity by rejecting self-pollen but allowing non-self-pollen for fertilization.The pear and other Rosaceae plants have the gametophytic self-incompatibility.The non-self S-RNase will be ubiquitined by SCF complex in pollen tube and then degrated by the 26S proteasome lead to normal fertilization,while the self-S-RNase could not be ubiquitined and the toxicity implemented by self S-RNase will lead to the death of pollen.Here we found a CULLIN1 and a SKP1 gene play a important role for pollen tube growth underlying non-self S-RNase;found 18 SFBBs could be potential pollen S determinants and explored the possible SCF complex.The following are our major results:1.In this study,57 CUL genes were isolated from the sequenced genomes of seven Rosaceae trees and were divided into three subfamilies according to phylogenetic analyses.Evolutionary analysis indicated that multiple duplication events play important role in the expansion of the CUL gene family in Rosaceae.The expression pattern shows that only 4 CUL genes are highly expressed in pollen and pollen tube in pear.Of these CUL genes,PbCUL1.C1 present the highest levels of expression in pollen,but the knockdown of PbCUL1.C1 by antisense oligonucleotide(ASO)did not influence pollen tube growth.Interestingly,the pollen tube treated by both the non-self S-RNase and ASO of PbCUL1.C1 were longer than that treated by unique non-self S-RNase,suggesting that the PbCUL1.C1 play an important role for pollen tube growth underlying non-self S-RNase in pear.2.73 SKP1 genes were isolated from the five Rosaceae fruit trees and were divided into three subfamilies.Multiple duplication events involved in the expansion of the SKP1 gene family in Rosaceae.Only 5 SKP1 genes(PbSK4,PbSK5,PbSK7,PbSK13 and PbSK14)are highly expressed in pollen.The knock of PbSK13 and PbSK14 by antisense oligonucleotide(ASO)did not influence pollen tube growth.However,the pollen tube treated by both the non-self S-RNase and ASO of PbSK13 were shorter than that treated by unique non-self S-RNase,meanwhile PbSK13 could interact with the SFBBs,suggesting that PbSK13 could assemble potential SCF complex to involve in the degration of non-self S-RNase.3.Identified 20,23,16 and 12 SFBBs from the S7,S17,S genotype of Bartlett and Pyrus betulaefolia.There are 18 SFBBs around S17-RNase linked with S17-RNase;this 18 SFBBs were all specifically expressed in pollen;the sequence analysis show that this SFBBs exhibits high sequence diversity.4.By Yeast two-hybrid assay and Firefly Luciferase Complementation Assay.The interaction between SFBBs and SKP1 genes were detected.As a result,11 interactions were found between 6 SFBBs(SFBB17-U1,SFBB17-U4,SFBB17-U13,SFBB17-D1,SFBB17-D2 and SFBB17-D4)and 6 SKP1 genes(PbSK3,PbSK7,PbSK9,PbSK10,PbSK13 and PbSK15).