Analysis of QTL DM4.1 for Downy Mildew Resistance in Cucumber Reveals Multiple subQTL: A Novel RLK as Candidate Gene for the Most Important subQTL

Abstract

One of the biggest problems in cucumber cultivation is cucurbit downy mildew (DM), caused by the obligate biotroph Pseudoperonospora cubensis. Whereas DM in cucumber was previously efficiently controlled by the dm-1 gene from Indian cucumber accession PI 197087, this resistance was broken by new DM strains, prompting the search for novel sources of resistance. A promising source of resistance is the wild cucumber accession PI 197088. It was previously shown that DM resistance in this genotype inherits polygenically. In this paper, we put the focus on one of the QTL, DM4.1 that is located on chromosome 4. QTL DM4.1 was shown to consist of three subQTL: DM4.1.1 affected pathogen-induced necrosis, DM4.1.2 was shown to have an additive effect on sporulation, and DM4.1.3 had a recessive effect on chlorosis as well as an effect on sporulation. Near-isogenic lines (NILs) were produced by introgressing the subQTLs into a susceptible cucumber line (HS279) with good horticultural traits. Transcriptomic analysis revealed that many genes in general, and defense pathway genes in particular, were differentially expressed in NIL DM4.1.1/.2 compared to NIL DM4.1.3 and the susceptible parent HS279. This indicates that the resistance from subQTL DM4.1.1 and/or subQTL DM4.1.2 likely involves defense signaling pathways, whereas resistance due to subQTL DM4.1.3 is more likely to be independent of known defense pathways. Based on fine-mapping data, we identified the RLK gene CsLRK10L2 as a likely candidate for subQTL DM4.1.2, as this gene was found to have a loss-of-function mutation in the susceptible parent HS279, and was strongly upregulated by P. cubensis inoculation in NIL DM4.1.1/.2. Heterologous expression of this gene triggered necrosis, providing further evidence that this gene is indeed causal for subQTL DM4.1.2.

Identification of Novel Loci and Candidate Genes for Cucumber Downy Mildew Resistance Using GWAS

“…A QTL interval usually contains many genes, and genetic mapping will help to narrow down the QTL interval. This has been demonstrated in the study of Berg et al [11], wherein three subQTLs were clustered in one QTL locus. Each subQTL contributes to DM resistance by limiting different infection processes of DM including chlorosis, sporulation, and DM-induced necrosis [11].…”

“…This has been demonstrated in the study of Berg et al [11], wherein three subQTLs were clustered in one QTL locus. Each subQTL contributes to DM resistance by limiting different infection processes of DM including chlorosis, sporulation, and DM-induced necrosis [11]. By developing and using near-isogenic lines, these subQTLs could be differentiated and causal genes underlying two subQTLs have been cloned [11,12].…”

“…3, since this mutation was not present in the CG lines (a subset of the resequenced 115 cucumber genome [ 13 ]) [ 12 ]. Further, the CsLRK10L2 gene underlying the resistance conferred by the DM4.1 QTL in PI 197088 [ 11 ] was located 1Mb away from the peak dmG4.2 . From the 32 CG lines that were identified carrying the PI 197088 allele [ 11 ], DI data were available for 27 lines ( Supplementary File 6 ).…”

“…Unfortunately, the mutant allele described in Berg et al [12] could not explain the DM resistance of dmG4.3, since this mutation was not present in the CG lines (a subset of the resequenced 115 cucumber genome [13]) [12]. Further, the CsLRK10L2 gene underlying the resistance conferred by the DM4.1 QTL in PI 197088 [11] was located 1Mb away from the peak dmG4.2. From the 32 CG lines that were identified carrying the PI 197088 allele [11], DI data were available for 27 For the locus dmG7.2, no SNPs with significant effect on DI could be identified.…”

“…Very recently, it was shown that the QTL DM4.1 identified in PI 197088 contains three subQTLs, with each contributing to chlorosis, sporulation, and necrosis, respectively. One cucumber receptor-like kinase (RLK) gene CsLRK10L2 is potentially the casual gene for subQTL DM4.1.2 contributing to sporulation [ 11 ]. Further, a loss-of-function mutation in the cucumber Amino Acid Permease 2A ( CsAAP2A gene) underlies the resistance conferred by subQTL4.1.3 through limiting the DM-induced chlorosis [ 12 ].…”