Cryptic Variation in Adaptive Phenotypes Revealed by PanspecificflcMutants

Abstract

The study of mutants is one of the best tools for understanding the genetic basis of phenotypes that contribute to adaptation. Oddly, mutant analyses are almost always restricted to single genetic backgrounds and findings therefore can not be easily generalized. A case in point is the key regulator of flowering,FLOWERING LOCUS C(FLC), which has been inferred to explain much of the flowering time variation inArabidopsis thaliana, yet mutants have been examined in very few backgrounds. We have previously established a set of panspecificflcmutants in 62 accessions ofA. thaliana(Ruffley et al. 2024). Here, we investigate how genetic background modulates mutant effects on flowering and vegetative traits, as well as on physiology and transcriptomes. Time to onset of flowering in the genome-editedflclines was reduced by up to 83%, but considerable variation remained. Genetic mapping showed that extremely early flowering in the absence ofFLCwas mostly explained by natural variation at the known FLC targetFT, with additional contribution from loci colocalizing withFLC. Prognostic sequence analyses of accessions did not suggest that extremely-early combinations of engineeredflcand naturalFTalleles would be deleterious, yet extremely early flowering accessions are not represented in the commonly used collections ofA. thalianaaccessions. To test whether this discrepancy could be due to sampling bias, we undertook a focused collection effort of wild populations in Southern Italy, which confirmed that extremely early flowering accessions exist in natural populations. Apart from its specific role in flowering time regulation,FLChas pleiotropic effects on other ecophysiological traits such as growth, and these were also dependent on the genetic background, which was further supported by transcriptomic comparisons. Together we conclude that the various roles ofFLChave greatly diversified in different genetic backgrounds. Our study provides a proof-of-concept on how analysis of panspecific mutants can reveal the true extent of genetic networks in which a focal gene participates in.