ABSTRACT More than total adiposity, the relative distribution of adipose tissue among central and peripheral depots is a critical determinant of Type 2 Diabetes (T2DM) and cardio metabolic disease risk. The goal of this proposal is to use novel mesenchymal progenitor cell derivation and single-cell clone genomic sequencing approaches to fully identify and characterize the diversity of adipocytes that compose these human depots and their developmental mechanisms. To this end, we will leverage exciting recent methods whereby we can generate large numbers of mesenchymal progenitor cells from human adipose tissue with minimal loss of multipotency. These cells differentiate into adipocytes that are similar to those from the depot of origin, and their transcriptomes reveal the existence of at least three types of “white” human adipocytes, as well as the thermogenic “beige/brite” type. We are now in a strong position to test the hypothesis that metabolically distinct human adipose depots, gluteal and abdominal, are composed of different adipocyte classes that develop from specific mesenchymal progenitor cells, and to provide full transcriptomic profiles of these adipocytes and their progenitors. Furthermore, it is known that diabetes affects multipotent progenitor cells, leading to impaired capacity to generate healthy adipocytes and repair tissue, further deterioration of insulin responsiveness. We will test the hypothesis that human T2DM alters mesenchymal progenitor diversity and determine how this defect leads to abnormal adipose tissue development in vivo. Our specific aims are: 1. To test the hypothesis that the different functional properties of human adipose tissue depots are due to intrinsic differences in their content of adipocyte subtypes derived from specific mesenchymal progenitor cells. 2: To test the hypothesis that T2DM preferentially impairs development of specific mesenchymal progenitor subsets and adipocytes derived from these cells, and 3: To define the physiological properties of adipocyte subtypes through tissue generation in vivo. These studies will provide a new high-resolution view of the cellular structure of human adipose tissue depots, of developmental mechanisms that lead to adipocyte subtypes, and insight into developmental alterations that contribute to T2DM physiopathology.