RNA-seq of in vitro differentiated human abdominal and gluteal adipocyte cell lines following doxycycline induced RSPO3-knockdown. RNA-seq of in vitro differentiated human abdominal and gluteal adipocyte cell lines following doxycycline induced RSPO3-knockdown

GWAS studies and our own work have identified RSPO3 as a gene modulating human body fat distribution. The GWAS signal at RSPO3 is coincident with an eQTL in mature adipocytes. To assess the effects of RSPO3 on abdominal and gluteal adipocyte biology, we undertook inducible RSPO3-knockdown in in vitro differentiated immortalized human abdominal and gluteal adipocyte cell lines (DFAT cells). Overall design: Abdominal and gluteal DFAT preadipocyte cells stably transduced with the tet-pLKO-puro-shRSPO3 (tet-shRSPO3) or tet-pLKO-puro-shCON (tet-shCON) lentiviral vectors were differentiated in standard adipogenic media. On day 13 of differentiation, cells were treated with doxycycline (final concentration of 0.05 µg ml-1), or vehicle, in hormone-free basal media for ~48 hours. On day 15, cells were harvested for RNA. Total RNA purification and on-column DNase I-treatment were performed using the RNeasy Mini kit (Qiagen). RNA-seq was performed on samples from three independent experiments. In addition to tet-shCON cells, vehicle-treated cells were used as controls.