Seifert Anja, Posern Guido
Institute for Physiological Chemistry, Medical Faculty, Martin Luther University Halle-Wittenberg, 06114, Halle (Saale), Germany.
Breast Cancer Res. 2017 Jun 7;19(1):68. doi: 10.1186/s13058-017-0860-3.
Myocardin-related transcription factors (MRTF) A and B link actin dynamics and mechanotransduction to gene expression. In mice, MRTF-A is involved in mammary gland differentiation, but its role in human mammary epithelial cells remains unclear.
Three-dimensional cultures of human mammary epithelial MCF10A cells were used to model acinar morphogenesis. Stable MRTF-A knockdown, MRTF-A/B rescue and MRTF-A/B overexpression was established to characterize the functional role during morphogenesis using confocal microscopy and expression analysis. Breast cancer patient databases were analyzed for MRTF-A expression.
We showed that a precise temporal control of MRTFs is required for normal morphogenesis of MCF10A mammary acini. MRTF transcriptional activity, but not their protein amounts, is transiently induced during 3D acini formation. MRTF-A knockdown dramatically reduces acini size and prevents lumen formation. These effects are rescued by re-expression of MRTF-A, and partially by MRTF-B. Conversely, overexpression of MRTF-A and MRTF-B increases acini size, resulting in irregular spheroids without lumen and defective apico-basal polarity. These phenotypes correlate with deregulated expression of cell cycle inhibitors p21/Waf1, p27/Kip1 and altered phosphorylation of retinoblastoma protein. In MRTF overexpressing spheroids, proliferation and apoptosis are simultaneously increased at late stages, whilst neither occurs in control acini. MRTFs interfere with anoikis of the inner cells and cause an integrin switch from α6 to α5, repression of E-cadherin and induction of mesenchymal markers vimentin, Snai2 and Zeb1. Moreover, MRTF-overexpressing spheroids are insensitive to alteration in matrix stiffness. In two breast cancer cohorts, high expression of MRTF-A and known target genes was associated with decreased patient survival.
MRTF-A is required for proliferation and formation of mammary acini from luminal epithelial cells. Conversely, elevated MRTF activity results in pre-malignant spheroid formation due to defective proliferation, polarity loss and epithelial-mesenchymal transition.
心肌相关转录因子（MRTF）A和B将肌动蛋白动力学和机械转导与基因表达联系起来。在小鼠中，MRTF-A参与乳腺分化，但其在人乳腺上皮细胞中的作用尚不清楚。
用人乳腺上皮MCF10A细胞的三维培养来模拟腺泡形态发生。通过共聚焦显微镜和表达分析，建立稳定的MRTF-A敲低、MRTF-A/B拯救和MRTF-A/B过表达模型，以表征形态发生过程中的功能作用。分析乳腺癌患者数据库中MRTF-A的表达情况。
我们发现，MCF10A乳腺腺泡的正常形态发生需要MRTF的精确时间控制。在三维腺泡形成过程中，MRTF的转录活性而非其蛋白量被短暂诱导。敲低MRTF-A会显著减小腺泡大小并阻止管腔形成。重新表达MRTF-A可挽救这些效应，MRTF-B可部分挽救。相反，过表达MRTF-A和MRTF-B会增加腺泡大小，导致无管腔的不规则球体以及顶端-基底极性缺陷。这些表型与细胞周期抑制剂p21/Waf1、p27/Kip1的表达失调以及视网膜母细胞瘤蛋白磷酸化改变相关。在过表达MRTF的球体中，晚期增殖和凋亡同时增加，而对照腺泡中均未发生。MRTF干扰内部细胞的失巢凋亡，导致整合素从α6转换为α5，抑制E-钙黏蛋白并诱导间充质标志物波形蛋白、Snai2和Zeb1。此外，过表达MRTF的球体对基质硬度的改变不敏感。在两个乳腺癌队列中，MRTF-A和已知靶基因的高表达与患者生存率降低相关。
MRTF-A是管腔上皮细胞增殖和形成乳腺腺泡所必需的。相反，MRTF活性升高会由于增殖缺陷、极性丧失和上皮-间质转化导致癌前球体形成。
