Functional analysis of Arabidopsis pollen coat proteins in the pollen-stigma interaction

Abstract

Sexual reproduction in Arabidopsis thaliana is a highly regulated process that is safeguarded by different molecular ‘checkpoints’. Successful reproduction relies on a complex network of interactions between male (pollen) and female reproductive cells and tissues (stigma) during pollination to pass through all the ‘checkpoints’. Arguably, the first ‘checkpoint’ – the molecular discrimination of pollen and establishment of its compatibility status at the pollen stigma interface is one of the most vital process in early stages of sexual reproduction. Previous work on Arabidopsis and Brassica have focused on characterisation of pollen-borne factors that regulate pollen-stigma compatibility. These work have established a paradigm that cross-talk between the PCP-B (pollen coat protein, class B) proteins and a pair of stigmatic receptors are the essential component for a compatible pollen to progress through the first molecular reproduction ‘checkpoints’. However, bilateral removal of these ligands and receptors did not incapacitate the mutant plant’s ability to accept compatible pollen, prompting further research into other pollen-borne and stigmatic compatibility factors. In this study, two pollen coat proteins, both belong to the family of rapid alkalinzation factor family, were characterised and determined to be novel pollen compatibility regulating factors. Meanwhile, a leucine-rich repeat receptor-kinase (LRR-RK) in the stigma was also characterised and is emerging as a novel stigmatic-borne compatibility regulating factor. To further understand the molecular mechanism of the compatibility pathway and to elucidate the roles of all the known components (including the roles of two novel RALFs and the novel LRR-RK), protein-protein interaction assays were carried out with heterologusly expressed pollen-borne compatibility determinants (PCP-Bs and the two novel RALF proteins) and total stigmatic plasma membrane proteins in native lipid
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nanodiscs solubilised by styrene maleic-acid copolymers. The characterisation works unveiled other potential candidates that may also function in pollen-stigma interaction. This project adds new molecular players to the current understanding on the mechanisms underlying intraspecific and interspecific compatibility in angiosperms, and provides clues for future studies in the same area of research.

Date of Award	15 Nov 2023
Original language	English
Awarding Institution	University of Bath
Supervisor	James Doughty (Supervisor), Volkan Cevik (Supervisor) & Paul Whitley (Supervisor)