Mutations in a Leucine-Rich Repeat Receptor-Like Kinase gene result in male sterility and reduction in the number and size of fruit warts in cucumber (Cucumis sativus L.).

Key message: Mutations in the CsEMS1 gene result in male sterility and reduced wart number and density. Male sterility and fruit wart formation are two significant agronomic characteristics in cucumber (Cucumis sativus), yet knowledge of our underlying genetics is limited. In this study, we identified an EMS-induced male sterility and few small warts mutant (msfsw). Histological observations revealed defects the absence of tapetum, meiotic aberration and impaired microspore formation in the anthers of the mutant. The mutant also exhibits a reduction in both the size and number of fruit spines and fruit tubercules. Genetic analysis revealed that a single recessive gene is responsible for the mutant phenotypes. BSA-Seq and fine genetic mapping mapped the msfsw locus to a 63.7 kb region with four predicted genes. Multiple lines of evidence support CsEMS1(CsaV3_3G016940) as the candidate for the mutant allele which encodes an LRR receptor-like kinase, and a non-synonymous SNP inside the exon of CsEMS1 is the causal polymorphisms for the mutant phenotypes. This function of CsEMS1 in determination of pollen fertility was confirmed with generation and characterization of multiple knockout mutations with CRISPR/Cas9 based gene editing. In the wild-type (WT) plants, CsEMS1 was highly expressed in male flowers. In the mutant, the expression level of CsEMS1, several tapetum identity-related genes, and trichome-related genes were all significantly reduced as compared with the wild-type. Protein-protein interaction assays revealed physical interactions between CsEMS1 and CsTPD1. Quantitation of endogenous phytohormones revealed a reduction in the ethylene precursor ACC in CsEMS1 knockout lines. This work identified an important role of CsEMS1 in anther and pollen development as well as fruit spine/wart development in cucumber.