Yang Ying, Stopka Tomás, Golestaneh Nady, Wang Yan, Wu Kongming, Li Anping, Chauhan Bharesh K, Gao Chun Y, Cveklová Kveta, Duncan Melinda K, Pestell Richard G, Chepelinsky Ana B, Skoultchi Arthur I, Cvekl Ales
Department of Ophthalmology and Visual Sciences, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
EMBO J. 2006 May 17;25(10):2107-18. doi: 10.1038/sj.emboj.7601114. Epub 2006 May 4.
Pax6 and c-Maf regulate multiple stages of mammalian lens development. Here, we identified novel distal control regions (DCRs) of the alphaA-crystallin gene, a marker of lens fiber cell differentiation induced by FGF-signaling. DCR1 stimulated reporter gene expression in primary lens explants treated with FGF2 linking FGF-signaling with alphaA-crystallin synthesis. A DCR1/alphaA-crystallin promoter (including DCR2) coupled with EGFP virtually recapitulated the expression pattern of alphaA-crystallin in lens epithelium and fibers. In contrast, the DCR3/alphaA/EGFP reporter was expressed only in 'late' lens fibers. Chromatin immunoprecipitations showed binding of Pax6 to DCR1 and the alphaA-crystallin promoter in lens chromatin and demonstrated that high levels of alphaA-crystallin expression correlate with increased binding of c-Maf and CREB to the promoter and of CREB to DCR3, a broad domain of histone H3K9-hyperacetylation extending from DCR1 to DCR3, and increased abundance of chromatin remodeling enzymes Brg1 and Snf2h at the alphaA-crystallin locus. Our data demonstrate a novel mechanism of Pax6, c-Maf and CREB function, through regulation of chromatin-remodeling enzymes, and suggest a multistage model for the activation of alphaA-crystallin during lens differentiation.
Pax6和c-Maf调控哺乳动物晶状体发育的多个阶段。在此，我们鉴定了αA-晶体蛋白基因的新型远端调控区域（DCRs），αA-晶体蛋白是由FGF信号诱导的晶状体纤维细胞分化的标志物。DCR1在用FGF2处理的原代晶状体外植体中刺激报告基因表达，将FGF信号与αA-晶体蛋白合成联系起来。一个与EGFP偶联的DCR1/αA-晶体蛋白启动子（包括DCR2）几乎重现了αA-晶体蛋白在晶状体上皮和纤维中的表达模式。相比之下，DCR3/αA/EGFP报告基因仅在“晚期”晶状体纤维中表达。染色质免疫沉淀显示Pax6与晶状体染色质中的DCR1和αA-晶体蛋白启动子结合，并证明αA-晶体蛋白的高表达水平与c-Maf和CREB与启动子的结合增加以及CREB与DCR3的结合增加相关，从DCR1延伸到DCR3的组蛋白H3K9高乙酰化的广泛区域，以及αA-晶体蛋白基因座处染色质重塑酶Brg1和Snf2h的丰度增加。我们的数据通过对染色质重塑酶的调控证明了Pax6、c-Maf和CREB功能的新机制，并提出了晶状体分化过程中αA-晶体蛋白激活的多阶段模型。
