A comprehensive fitness landscape model reveals the evolutionary history and future evolvability of eukaryotic cis-regulatory DNA sequences

Mutations in non-coding cis-regulatory DNA sequences can alter gene expression, organismal phenotype, and fitness. Fitness landscapes, which map DNA sequence to organismal fitness, are a long-standing goal in biology, but have remained elusive because it is challenging to generalize accurately to the vast space of possible sequences using models built on measurements from a limited number of endogenous regulatory sequences. Here, we construct a sequence-to-expression model for such a landscape and use it to decipher principles of cis-regulatory evolution. Using tens of millions of randomly sampled promoter DNA sequences and their measured expression levels in the yeast Sacccharomyces cerevisiae, we construct a deep transformer neural network model that generalizes with exceptional accuracy, and enables sequence design for gene expression engineering. Using our model, we predict and experimentally validate expression divergence under random genetic drift and strong selection weak mutation regimes, show that conflicting expression objectives in different environments constrain expression adaptation, and find that stabilizing selection on gene expression leads to the moderation of regulatory complexity. We present an approach for detecting selective constraint on gene expression using our model and natural sequence variation, and validate it using observed cis-regulatory diversity across 1,011 yeast strains, cross-species RNA-seq from three different clades, and measured expression-to-fitness curves. Finally, we develop a characterization of regulatory evolvability, use it to visualize fitness landscapes in two dimensions, discover evolvability archetypes, quantify the mutational robustness of individual sequences and highlight the mutational robustness of extant natural regulatory sequence populations. Our work provides a general framework that addresses key questions in the evolution of cis-regulatory sequences.