Altered metabolic responses to stress render darier disease keratinocytes prone to DNA repair abnormalities

Summary: Abstract Body: Despite a known role in skin health, calcium dynamics remain underexplored in terms of therapeutics. The heritable acantholytic disorder, Darier disease (DD), arises from well-studied heterozygous variants of the ER calcium pump, SERCA2. Like prior studies, we measured aberrant calcium levels in DD keratinocytes (KCs) but pursued a novel hypothesis. As stress-induced flares characterize DD, we hypothesized that calcium dysregulation could hamper the metabolic response to stress. To test this idea, metabolic profiling of 3 DD patient isolates was conducted. When control KCs were exposed to an oxidative stressor, a set of 44 metabolites showed strong positive Pearson correlations (rho= 0.74 +/- 0.15; mean+/- sd), matched by a negatively correlated group of 27 metabolites (rho= -0.48 +/- 0.23 sd). A coordinated loss of glycolytic products and a surge of pentose phosphate pathway (PPP) metabolites drove these metrics. The correlations collapsed in DD cells (rho= 0.27+/-0.42 and -0.08 +/- 0.31, respectively). DD cells, then, failed to shunt intermediates from glycolysis to the parallel, PPP. Stressed cells normally activate the PPP to replenish antioxidants and produce nucleotides for DNA repair. Western blotting revealed elevated γ-H2AX levels in DD cells, suggesting chronic activation of the DNA repair system and the likelihood of heightened genetic variability. Indeed, publicly available RNAseq reads for 4 control and 4 DD skin samples deviated from the reference genome once every 5681+/-1922 and 862+/-127 exon-mapped bases (p=0.002), respectively. These observations provide a unique perspective on the origin of somatic SERCA2 variants reported to cause type 2 mosaicism in DD and a recently published link to carcinoma. Furthermore, the muted metabolic capacity for protecting genomic integrity highlights a need to consider PPP activation and genoprotective agents in DD therapeutic development. Robert Harmon², Erin McCarthy², Jenny Pokorny², Lisa M. Godsel², Ziyou Ren², Eran Cohen Barak¹, Roni Dodiuk-Gad¹, Amy Paller², Kathleen Green² 1. Technion Israel Institute of Technology, Haifa, Haifa District, Israel. 2. Northwestern University, Evanston, IL, United States. Genetic Disease, Gene Regulation, Gene Therapy & Epigenetics