Homeostatic levels of SRC-2 and SRC-3 promote early human adipogenesis

The related coactivators SRC-2 and SRC-3 interact with peroxisome proliferator activated receptor gamma (PPAR gamma) to coordinate transcriptional circuits to promote adipogenesis. To identify potential coactivator redundancy during human adipogenesis at single cell resolution, we used high content analysis to quantify links between PPAR gamma, SRC-2, SRC-3, and lipogenesis. Because we detected robust increases and significant cell cell heterogeneity in PPAR gamma and lipogenesis, without changes in SRC-2 or SRC-3, we hypothesized that permissive coregulator levels comprise a necessary adipogenic equilibrium. We probed this equilibrium by down-regulating SRC-2 and SRC-3 while simultaneously quantifying PPAR gamma. Individual or joint knockdown equally inhibits lipid accumulation by preventing lipogenic gene engagement, without affecting PPAR gamma protein levels. Supporting dominant, pro-adipogenic roles for SRC-2 and SRC-3, SRC-1 knockdown does not affect adipogenesis. SRC-2 and SRC-3 knockdown increases the proportion of cells in a PPAR gamma(hi)/lipid(lo) state while increasing phospho-PPAR gamma-S114, an inhibitor of PPAR gamma transcriptional activity and adipogenesis. Together, we demonstrate that SRC-2 and SRC-3 concomitantly promote human adipocyte differentiation by attenuating phospho-PPAR gamma-S114 and modulating PPAR gamma cellular heterogeneity.