Natural variation in Arabidopsis responses toPlasmodiophora brassicaereveals an essential role for RPB1

Abstract

ABSTRACTDespite the identification of clubroot resistance genes in various Brassica crops our understanding of the genetic basis of immunity toPlasmodiophora brassicaeinfection in the model plantArabidopsis thalianaremains limited. To address this issue we performed a screen of 142 natural accessions and identified 11 clubroot resistant Arabidopsis lines. Genome wide association analysis identified several genetic loci significantly linked with resistance. Three genes from two of these loci were targeted for deletion by CRISPR/Cas9 mutation in resistant accessions Est-1 and Uod-1. Deletion ofResistance to Plasmodiophora brassicae 1(RPB1) rendered both lines susceptible to theP. brassicaepathotype P1+. Further analysis ofrpb1knock-out Est-1 and Uod-1 lines showed that the RPB1 protein is required for activation of downstream defence responses, such as the expression of phytoalexin biosynthesis geneCYP71A13. RPB1 has no known functional domains or homology to previously characterised proteins. The clubroot susceptible Arabidopsis accession Col-0 lacks a functionalRPB1gene; when Col-0 is transformed withRPB1expression driven by its native promoter it is capable of activatingRPB1expression in response to infection but this is not sufficient to confer resistance. Constitutive over-expression ofRPB1in Col-0 leads to drastically reduced growth and activation of stress-responsive genes. Furthermore, we found that transient expression ofRPB1inNicotiana tabacuminduced programmed cell death in leaves. We conclude that RPB1 is a critical component of the defence response toP. brassicaeinfection in Arabidopsis, acting downstream of pathogen recognition but required for the elaboration of effective resistance.