GLUCOSE BINDS RBPs TO REGULATE EPIDERMAL HOMEOSTASIS PROJECT SUMMARY/ABSTRACT One in four Americans are affected by skin disease, underscoring the importance of understanding the mechanisms underlying epidermal homeostasis. Our laboratory recently found that increases in intracellular glucose concentrations are essential for differentiation in the epidermis, independent of glucose’s metabolic function. In this context, glucose acts to promote or dissociate specific protein multimers. RNA-binding proteins (RBPs) are required for the control of keratinocyte proliferation and differentiation. Using CRISPR-FlowFISH and glucose pull-down MS techniques, we have identified RBPs as the primary glucose binding proteins that control epidermal differentiation. Among these RBPs, DDX21 was validated as a key regulator of glucose-mediated RNA and protein interactions needed to maintain normal skin homeostasis. Additionally, we have discovered other RBPs that bind glucose and are essential for epidermal differentiation. The objective of this K01 application is to characterize these essential glucose binding proteins in epidermis. This research will provide further insights into the non-metabolic roles of glucose in epidermal differentiation and will uncover mechanisms of RBP function that are regulated by direct glucose binding. In Aim I, we will define the mechanism for glucose-mediated regulation of DDX50 in facilitating epidermal differentiation. Our preliminary data supports the notion that glucose plays a critical role in regulating DDX50 in this context. Specifically, in Aim IA, we will assess the ability of glucose to enable the formation of the DDX50- RNA-STAU1 complex and investigate how the function of the complex is regulated by glucose. In Aim IB, we will explore alternative models to understand how glucose regulates DDX50's function in epidermal differentiation. In Aim II, we will test our hypothesis that glucose plays a role in directing NSUN2 and other RBPs to control epidermal homeostasis. Our preliminary data suggests that glucose facilitates S-adenosyl methionine (SAM) binding to NSUN2, which promotes epidermal differentiation. We have also identified 51 additional RBPs with glucose-binding affinity that are also essential for epidermal differentiation. In Aim IIA, we will focus on testing the specific role of glucose in regulating NSUN2 in epidermal differentiation. In Aim IIB, we will evaluate the role of glucose in regulating additional RBPs involved in epidermal differentiation. My background is in proteomics, molecular biology, RBP biology and skin differentiation. Over the course of the training period and within the environment of Stanford’s Department of Dermatology, I plan to acquire expertise in (1) statistics, (2) bioinformatics, (3) genomics. This training will enable me to develop a more comprehensive understanding of the mechanisms underlying glucose regulation in epithelial tissue, which is a key area of interest in my rese...