Investigating a role forPUB17andPUB16in the self-incompatibility pathway in transgenicArabidopsis thaliana

Abstract

AbstractIn Brassicaceae self-incompatibility (SI), self-pollen rejection is initiated by theS-haplotype specific interactions between the pollen SCR/SP11 ligand and the stigma S Receptor kinase (SRK). InBrassicaSI, a member of the Plant U-Box (PUB) E3 ubiquitin ligases, ARC1, is then activated by SRK in this stigma and cellular events downstream of this cause SI pollen rejection by inhibiting pollen hydration and pollen tube growth. During the transition to selfing,Arabidopsis thalianalost the SI components,SCR, SRK, andARC1. However, this trait can be reintroduced intoA. thalianaby adding back functional copies of these genes from closely related SI species. BothSCRandSRKare required for this, though the degree of SI pollen rejection varies between accessions, andARC1is not always needed to produce a strong SI response. ForA. thalianaC24, only transforming withA. lyrata SCRandSRKconfers a strong SI trait, and so here we investigated if ARC1-related PUBs were involved in the SI pathway. Two close ARC1 paralogs,PUB17andPUB16, were selected, and CRISPR/Cas9 technology was used to generatepub17andpub16mutations in the C24 accession. These mutants were then crossed into a transgenicA. thalianaSI-C24 line and their potential impact on SI pollen rejection was investigated. Overall, we did not observe any significant differences to implicatePUB17andPUB16functioning in the transgenicA. thalianaSI-C24 stigma to reject SI pollen.