Main candidate genes within PEK are a cluster of TNLs receptors
To identify candidate genes for the HR phenotype, we checked the annotations within the PEK locus in the B. nigra reference genome C2. The region contains 11 genes with many duplicated loci such as three B. nigra homologs of a methionine aminopeptidase 1D (MAP1D, AT4G3700), three homologs of an unknown membrane protein (AT4G37030) and three TIR-NBS-LRR. A fourth TIR-NBS-LRR (TNL) is present just outside marker M28. Based on gene annotations, the four TNLs are the only genes that we could associate to known plant immunity functions (Cui, Tsuda, & Parker, 2015). Thus, they represent good candidate genes for the HR phenotype. Further, we used the sequencing data from the BSA-seq to study the variation within the PEK locus. In total we identified 785 variants (SNPs and InDels) within each of the eleven genes, but we could not pinpoint yet putative casual variants associated to the trait (Supporting information: Table S6).
Finally, we investigated whether the PEK locus overlaps with loci that were previously associated with P. brassicae egg-induced cell death in A. thaliana (Groux et al., 2021) and B. rapa(Bassetti et al., 2022). Overall, PEK appeared syntenic to the bottom of A. thaliana chromosome 4 (Supporting information: Fig. S7a) and to the center of B. rapa chromosome A08 (Supporting information: Fig. S7b). None of these regions were previously associated with butterfly egg-induced HR, as genes AtLecRK-I.1 (AT3G45330) and AtGLR2.7 (AT2G29120) located on A. thalianachromosomes 2 and 3 (Groux, 2019; Groux et al., 2021), while B. rapa QTLs Pbc1-3 located on chromosomes A02, A03 and A06 (Bassetti et al., 2022). In conclusion, the PEK locus here identified in B. nigra represents a novel genomic region associated with a butterfly egg-induced HR which results in egg-killing.