The Scarabaeinae, or true dung beetles, are a hyper-diverse clade of insects of ecological, evolutionary, and agricultural significance and have long served as informative models of evolutionary ecology and development. Perhaps the most conspicuous of their unique traits are head horns, novel structures that serve as secondary sexual weapons, exhibit extraordinary developmental plasticity, and have fueled one of the most dramatic morphological radiations in the animal kingdom. In this study, we investigate the evolutionary basis for dung beetle traits - including horns - via comparative genomic and developmental assays. We present chromosome-level genome assemblies of three dung beetle species in the species-rich Onthophagini tribe (> 2500 extant species) including Onthophagus taurus, Onthophagus sagittarius, and Digitonthophagus gazella. Contrasting these assemblies with seven other species across the order Coleoptera identifies rapidly evolving gene families associated with metabolic regulation of developmental plasticity and metamorphosis. Intraspecific comparisons of chromatin accessibility in developing head horns of O. taurus identify distinct cis-regulatory architectures underlying sex- and nutrition-responsive development of this novel trait, including a large proportion of recently evolved regulatory elements sensitive to horn morph determination. Binding motifs of diverse developmental transcription factors are enriched in these nutrition-responsive open chromatin regions, including the early embryonic patterning gene twist. Using RNA interference (RNAi), we show twist has been co-opted into the beetle horn regulatory network to mediate differential horn morphogenesis in alternate male morphs via its interactions with nutrition-sensitive DNA-binding sites, highlighting the utility of this approach in identifying new developmental regulators of morphological evolution. These results demonstrate gene networks are highly evolvable transducers of environmental and genetic signals critical for the formation and diversification of developmental traits, established in part by condition-responsive chromatin accessibility. Further, this work provides new reference-quality genome assemblies of three dung beetles that will bolster future developmental, ecological, and evolutionary studies of this insect group.