Genetic and genomic analysis of electron transport chain Complex II functions in mammalian skin development - Project Summary The electron transport chain (ETC) is central to cellular metabolism, facilitating ATP synthesis through the establishment of an electromotive proton gradient across the inner mitochondrial membrane. Pioneering work in mitochondrial biology has revealed an additional role for the ETC in cellular signaling, both through the generation of reactive oxygen species (ROS) and through modulation of epigenetic and post-translational modifying enzyme activity by TCA cycle metabolites. While considerable effort has been made towards characterizing the effects of ETC dysfunction in tumorigenesis and human disorders of central metabolism, the role of individual ETC complexes during tissue morphogenesis remains largely unknown. Though previous work by our lab has shown that epidermal loss of ETC Complex III results in embryonic lethality and severe barrier defects through impaired epidermal differentiation, whether this defect reflects a requirement for Complex III activity, or a specific dependency on Complex I or II remains unclear. To answer this question, we have generated two novel mouse models with epithelial-specific deletions of Ndufs2, a critical Complex I component, and Sdhd, a crucial subunit of Complex II. Intriguingly, our preliminary data has shown that Ndufs2 is largely dispensable for normal hair and skin development, while Sdhd deletion results in numerous morphological defects, comprising a patchy hair coat, bumpy, lesioned skin, keratin plugging of the hair canal, and the expansion of infundibular and sebaceous gland populations. Loss of Sdhd is associated with an increase in Myc expression within the hair follicle, suggesting that these malformations are driven by an abnormal Myc-driven transit amplifying program within the upper hair follicle and sebaceous gland. These results contrast with a recent study by our collaborator, Navdeep Chandel, which showed that Complex I, but not Complex II, was essential for normal lung development, suggesting that individual respiratory complexes regulate epithelial development in a lineage-specific manner. I hypothesize that loss of a functional Complex II drives alterations in transcriptional programs, chromatin state, and gene expression via inhibition of a- ketoglutarate-dependent epigenetic modifiers, resulting in compromised epithelial stem cell fate specification, lineage formation, and differentiation. Here, I propose to (1) determine the function of Complex II in epidermal fate specification and skin morphogenesis and (2) investigate the mechanisms through which Complex II controls the epithelial cell transcriptome. Experiments proposed in Aim 1 will focus on conducting a comprehensive phenotypic, transcriptomic, and metabolomic analysis of Sdhd cKO mice. Aim 2 focuses on the analysis of epigenetic perturbations induced by loss of Complex II function through Cut&Run sequencing as well as single cell multiomic identification of candidate transcriptional regulators driving the response to Sdhd deletion in epidermal lineages. My results will provide important new insights into the role of the ETC in tissue morphogenesis, with implications for the study of tumorigenesis, stem cell biology, and aging.
