ABSTRACT
Global climate change has the potential to devastate biological systems as organisms are exposed to novel temperature regimes. Increases in annual mean temperature have been accompanied by disproportionate rates of change in temperature across seasons, and winter is the season warming most rapidly. Yet, to our knowledge, no research has characterized the direct effects of winter warming on the biology of overwintering organisms. Here, we investigated the effects of winter warming stress on internal freezing temperatures (supercooling points) and metabolome profiles of diapausing Pieris rapae butterfly pupae. We show that after acute and chronic winter warming exposure, pupae had higher supercooling points and significant changes in metabolite abundances across the entire metabolome. Notably, there were warming-induced shifts in key biochemical pathways that likely support energy metabolism and cryoprotection. These physiological responses suggest that winter warming will threaten the survival of overwintering P. rapae pupae, and by extension winter warming may pose threats to other species that overwinter. Furthermore, we found evidence of local adaptation of supercooling in P. rapae, as we observed significantly lower supercooling points in Vermont individuals relative to North Carolina individuals. Moving forward, future research should focus on species-wide responses to winter warming events, particularly in the context of local warming patterns, to better predict how populations may differentially respond to changes in winter thermal environments.