Pregnancy-Induced Noncoding RNA (PINC) Associates with Polycomb Repressive Complex 2 and Regulates Mammary Epithelial Differentiation
(A–C) After 24 (A) and 72 (B) hrs of hormone induction, LeGO-1.0/1.6 cells show reduced levels of Csn2, Wap, and Ltf expression by qPCR. Target genes were normalized to Gapdh and set relative to LeGO-GFP levels. Data are presented as mean ± SEM (n = 6). (C) Overexpression of mPINC also reduces formation of domes. Domes were counted at 48 hrs post-hormone treatment from nine 20× fields/experiment and data represent mean ± SEM set relative to the dome number in the LeGO-GFP control group (n = 6). (D–F) After 24 (D) and 72 (E) hrs of hormone induction, LeGO-DCR cells show increased levels of Ltf (at 24 hrs) and Csn2, but not Wap, expression compared to LeGO-GFP cells. Target genes were normalized to Gapdh and set relative to LeGO-GFP levels. Data are presented as mean ± SEM (n = 6). (F) Overexpression of the DCR mutant enhances dome formation compared to the control. Experiment was performed and analyzed as described in panel C. (G) RT-PCR shows mPINC overexpressed transcript in LeGO-1.0/1.6 and LeGO-DCR HC11 cells compared to endogenous levels in LeGO-GFP cells. (H) qPCR shows mPINC overexpressed levels are high and fairly equivalent relative to mPINC endogenous levels in MECs purified from virgin mammary gland (V-MECs). Target gene was normalized to Gapdh. Data are presented as mean ± SEM for three independent experiments.