Integration of GWAS and TWAS to elucidate the genetic architecture of natural variation for leaf cuticular conductance in maize

The cuticle, a hydrophobic layer of cutin and waxes synthesized by plant epidermal cells, is the major barrier to water loss when stomata are closed. Dissecting the genetic architecture of natural variation for maize leaf cuticular conductance (gc) is important for identifying genes relevant to improving crop productivity in drought-prone environments. To this end, we performed an integrated genome- and transcriptome-wide association study (GWAS/TWAS) to identify candidate genes putatively regulating variation in leaf gc. Of the 22 plausible candidate genes identified, five were predicted to be involved in cuticle precursor biosynthesis and export, two in cell wall modification, nine in intracellular membrane trafficking, and seven in the regulation of cuticle development. A gene encoding an INCREASED SALT TOLERANCE1-LIKE1 (ISTL1) protein putatively involved in intracellular protein and membrane trafficking was identified in GWAS and TWAS as the strongest candidate causal gene. A set of maize nested near-isogenic lines that harbor the ISTL1 genomic region from eight donor parents were evaluated for gc, confirming the association between gc and ISTL1 in a haplotype-based association analysis. The findings of this study provide novel insights into the role of regulatory variants in the development of the maize leaf cuticle, and will ultimately assist breeders to develop drought-tolerant maize for target environments.