Department of Integrative Biosciences, College of Veterinary Medicine, Texas A&M University, College Station, TX 77843-4458, USA.
Development. 2010 Mar;137(6):945-52. doi: 10.1242/dev.041657. Epub 2010 Feb 11.
We have previously demonstrated that the bHLH/PAS transcription factor, singleminded 2s (Sim2s), is required for proper mammary ductal morphogenesis and luminal epithelial differentiation. Furthermore, loss of Sim2s in breast cancer cells resulted in downregulation of epithelial markers and acquisition of a basal-like phenotype. The objective of this study was to further define the role of Sim2s in mammary differentiation. We found that Sim2s is developmentally regulated throughout mammary gland development with highest expression during lactation. Mammary glands from nulliparous mice expressing Sim2s driven by the mouse mammary tumor virus (MMTV) long terminal repeat (LTR) promoter were morphologically indistinguishable from wild-type mice but displayed hallmarks of precocious lactogenic differentiation. These included elevated expression of the milk protein genes Wap and Csn2, and apical localization of the lactation marker Npt2b. Consistent with the in vivo results, Sim2s enhanced prolactin-mediated Csn2 expression in HC11 and CIT3 mouse mammary epithelial cells, and downregulation of Sim2s by shRNA in HC11 cells inhibited Csn2 expression. Chromatin immunoprecipitation (ChIP) analyses of the Csn2 gene found that Sim2s associates with the Csn2 promoter and re-ChIP experiments showed that Sim2s interacted with the RNA II polymerase (RNAPII) complex. Together, these data demonstrate, for the first time, that Sim2s is required for establishing and maintaining mammary gland differentiation.
我们之前已经证明，bHLH/PAS 转录因子单倍型 2s（Sim2s）对于乳腺导管形态发生和腔上皮分化是必需的。此外，乳腺癌细胞中 Sim2s 的缺失导致上皮标志物下调和获得基底样表型。本研究的目的是进一步定义 Sim2s 在乳腺分化中的作用。我们发现 Sim2s 在乳腺发育过程中具有发育调节作用，在哺乳期表达最高。由小鼠乳腺肿瘤病毒（MMTV）长末端重复（LTR）启动子驱动表达 Sim2s 的未产小鼠的乳腺在形态上与野生型小鼠无法区分，但表现出早熟泌乳分化的特征。这包括乳蛋白基因 Wap 和 Csn2 的表达升高，以及泌乳标志物 Npt2b 的顶端定位。与体内结果一致，Sim2s 增强了催乳素介导的 HC11 和 CIT3 小鼠乳腺上皮细胞中 Csn2 的表达，而 shRNA 下调 HC11 细胞中的 Sim2s 抑制了 Csn2 的表达。CSN2 基因的染色质免疫沉淀（ChIP）分析发现 Sim2s 与 CSN2 启动子结合，重新 ChIP 实验表明 Sim2s 与 RNA II 聚合酶（RNAPII）复合物相互作用。总之，这些数据首次表明 Sim2s 对于建立和维持乳腺分化是必需的。
