Repression of chondrogenesis through binding of notch signaling proteins HES-1 and HEY-1 to N-box domains in the COL2A1 enhancer site.
Repression of chondrogenesis through binding of notch signaling proteins HES-1 and HEY-1 to N-box domains in the COL2A1 enhancer site.
复制标题
DOI：
10.1002/art.23730
复制
发表时间：
2008-09
影响因子：
--
通讯作者：
Lotz, Martin K.
中科院分区：
文献类型：
--
作者：
Grogan, Shawn P.;Olee, Tsaiwei;Hiraoka, Koji;Lotz, Martin K.
Notch 信号传导参与抑制间充质干细胞 (MSC) 软骨形成分化。这种抑制的机制以及如何调节它以允许软骨形成尚未阐明。在 MSC 沉淀培养物中，通过蛋白质印迹法监测人软骨形成过程中的 Notch 胞内结构域 (NICD) 蛋白水平。通过转染编码NICD、毛状和分裂1增强子(Hes-1)以及毛状和分裂相关增强子2(HERP-2/Hey-1)的质粒，实现了Notch信号成分的过表达及其对软骨形成的影响。通过定量 PCR 监测 Col2a1 和聚集蛋白聚糖的表达。利用染色质免疫沉淀 (chIP) 来测试 Hes-1 和 Hey-1 是否结合 Col2a1 内含子 1 中推定的 N-box 结构域。 hMSC 软骨形成过程中 NICD 蛋白水平降低，这是由 TGFβ3 介导的。 Col2a1 基因表达在 NICD（2 倍）、Hes-1（3 倍）和 Hey-1（80 倍）过表达后受到抑制。 Hey-1 将聚集蛋白聚糖的表达抑制了 10 倍，而 NICD 和 Hes-1 则没有效果。 ChIP 研究表明，内源 Hes-1 和 Hey-1 与两个假定的 N-box 结构域结合，该结构域邻近位于 Col2a1 内含子 1 的 Sox9 增强子结合位点，并作为该位点的一部分。 Hes-1 共阻遏蛋白转导蛋白样增强子 (TLE) 在软骨分化过程中和 TGFβ3 治疗后被替换。这些结果揭示了 Notch 信号传导抑制基因表达的新机制，并进一步明确了 TGFβ 促进软骨形成分化的作用。
Notch signaling is implicated in repressing mesenchymal stem cell (MSC) chondrogenic differentiation. The mechanism of this repression and how this is modulated to permit chondrogenesis has not been elucidated. Notch intracellular domain (NICD) protein levels were monitored via western blot throughout chondrogenic differentiation of human in MSC pellet cultures. Overexpression of Notch signaling components and their effect on chondrogenesis was achieved by transfecting plasmids coding for NICD, hairy and enhancer of split 1 (Hes-1) and hairy and enhancer of split related-2 (HERP-2/Hey-1). Col2a1 and aggrecan expression was monitored via quantitative PCR. Chromatin immunoprecipitation (chIP) was utilized to test whether Hes-1 and Hey-1 bind putative N-box domains in intron 1 of Col2a1. NICD protein levels were reduced during chondrogenesis of hMSC, which was mediated by TGFβ3. Col2a1 gene expression was repressed following overexpression of NICD (2-fold), Hes-1 (3-fold) and markedly by Hey-1 (80-fold). Hey-1 repressed aggrecan expression 10-fold, while NICD and Hes-1 had no effect. chIP studies show that endogenous Hes-1 and Hey-1 bind to two putative N-box domains adjacent to and as part of the Sox9 enhancer binding site located in intron 1 of Col2a1. The Hes-1 co-repressor protein transducin like enhancer (TLE) was displaced during chondrogenic differentiation and following TGFβ3 treatment. These results reveal novel mechanisms by which Notch signaling represses gene expression and further define the role of TGFβ to promote chondrogenic differentiation.
登录
查看更多内容
影响因子：
64.5
作者：
Ju, BG;Solum, D;Rosenfeld, MG
通讯作者：
Rosenfeld, MG
影响因子：
4.6
作者：
Kageyama, Ryoichiro;Ohtsuka, Toshiyuki;Kobayashi, Taeko
通讯作者：
Kobayashi, Taeko
影响因子：
4.8
作者：
Kou, I;Ikegawa, S
通讯作者：
Ikegawa, S
影响因子：
--
作者：
Barbero, A;Ploegert, S;Martin, I
通讯作者：
Martin, I
影响因子：
64.8
作者：
JARRIAULT, S;BROU, C;ISRAEL, A
通讯作者：
ISRAEL, A