Pollen-pistil Interactions in the Genus Nicotiana

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The goal of this project is to identify and characterize factorsthat contribute to S-allele-specific pollen recognition in Nicotiana alata.In this species, the S-locus controls intraspecific compatibility. Pollenis rejected when the S-allele in the pollen is the same as one of theS-alleles expressed in the pistil. S-RNases control the specificity ofpollen rejection, but other factors contribute to S-allele-specific pollenrecognition in other ways. Our main objective is to identify andcharacterize these non-S-RNase factors. HT-protein is one such factor that appears to be an unstablecomponent of the stylar extracellular matrix. Antisense suppression of HTprevents S-allele-specific pollen rejection, but its precise function isunknown. We propose experiments to determine the abundance of HT-proteinduring style development, and why it is so unstable. We have found that twospecific stylar proteins bind to S-RNase in vitro. One is a previouslycharacterized protein, NaTTS, that is implicated in growth of compatiblepollen tubes. The second, designated p11, has not been well characterized.NaTTS and p11 are putative non-S-RNase factors that may be implicated inboth pollen rejection and in supporting compatible pollen tube growth. Gelfiltration and protein blot analysis are proposed to determine the molecularweight and composition of NaTTS/S-RNase/p11 complexes. NaTTS and p11 willbe also expressed in bacteria and then used to test for interactions invitro. Antisense methods will be used to determine whether p11 affectscompatible or incompatible pollinations in vivo. To identify newnon-S-RNase factors, we have generated populations that segregate for theability to perform S-allele-specific pollen rejection. cDNA AFLP will beused to screen for new sequences associated with pollen rejection phenotype. The impact of this research will be to provide a betterunderstanding of the genetic mechanisms for pollen recognition andrejection. This is of practical significance to biotechnologists who wishto manipulate plant breeding behavior.