Genome-wide association studies revealed association between LePC4 gene and crude protein content in shiitake mushroom, Lentinula edodes.

Lentinula edodes, the most widely cultivated edible mushroom worldwide, is of considerable significance for nutritional health and commercial applications owing to its relatively high crude protein content (CPC). However, the genetic mechanisms underlying the regulation of CPC in edible mushrooms remain poorly understood. In this study, genome-wide association studies (GWAS) identified twenty single-nucleotide polymorphism (SNP) loci significantly associated with CPC in L. edodes, implicating 31 candidate genes. Integrating GWAS results with comparative transcriptomic analysis of two L. edodes strains exhibiting markedly divergent CPC levels, we prioritized LePC4 as a key candidate gene. Conserved domain analysis indicated that LePC4 encodes a DUF5910-containing protein, suggesting a possible relationship with apoptosis-related protein. Subsequent functional characterization via both overexpression and RNA interference (RNAi) showed that interference of LePC4 significantly increased CPC in transformed mycelia by 13.06% - 17.11%, accompanied by elevated relative abundances of albumin (7.41% - 38.73%) and globulin (13.78% - 70.77%). Notably, LePC4 modulation did not adversely affect mycelial growth under the tested culture conditions. These findings suggest that LePC4 is involved in CPC variation in L. edodes and provide useful information for the development of high-protein shiitake germplasm.