Requirement of TCTG(G/C) Direct Repeats and Overlapping GATA Site for Maintaining the Cardiac-Specific Expression of Cardiac troponin T in Developing and Adult Mice.
Requirement of TCTG(G/C) Direct Repeats and Overlapping GATA Site for Maintaining the Cardiac-Specific Expression of Cardiac troponin T in Developing and Adult Mice.
复制标题
DOI：
10.1002/ar.20772
复制
发表时间：
2008-12
影响因子：
2
通讯作者：
Lin, Jim Jung-Ching
中科院分区：
文献类型：
--
作者：
Harlan, Shannon M.;Reiter, Rebecca S.;Sigmund, Curt D.;Lin, Jenny Li-Chun;Lin, Jim Jung-Ching
大鼠心肌肌钙蛋白T(CTnT)的心脏特异启动子−497bp含有两个相似的模块D和F，每个模块都有TCTG(G/C)直接重复序列和富含A/T的位点。为了鉴定心脏特异性的关键顺式元件，使用只含有模块F的−249BP启动子及其定点突变来产生转基因小鼠。−249BP启动子的转基因表达仍然是心脏特异的，尽管表达水平很低且不均匀。转基因的不均匀表达模式与HMGB1的差异表达相吻合，HMGB1似乎是心脏中HMGB家族蛋白的主要形式。染色质免疫沉淀分析表明，HMGB1与cTnT启动子的A/T富含/MEF2样位点结合。携带−249BP启动子的小鼠，其点突变扰乱了直接重复序列，在心脏和大脑中的异位表达水平较低。在发育中的四肢和头部也观察到外源基因的异位表达。这些结果表明，直接重复序列在确定心脏特异性方面具有重要作用。此外，携带突变启动子的小鼠同时扰乱了直接重复序列和重叠的GATA位点，在测试的任何组织中都无法表达转基因。因此，直接重复和重叠的GATA位点对GATA的表达水平和心脏特异性至关重要。F模块控制一个级别的心脏特异性。为了获得统一的高水平的心脏特异表达，还需要上游元件(−497bp到−250bp)，可能通过D增强子模块以及NKX2.5和GATA位点的组合。
The cardiac-specific −497bp promoter of rat cardiac troponin T (cTnT) contains two similar modules, D and F, each of which possesses TCTG(G/C) direct repeats and A/T-rich sites. To identify cis-elements critical for cardiac specificity, a −249bp promoter containing only module F and its site-directed mutations were used to generate transgenic mice. Transgene expression of the −249bp promoter remained cardiac-specific, despite low and non-uniform expression. The non-uniform expression pattern of the transgene coincided with differential expression of HMGB1, which appeared to be the predominant form of HMGB family proteins in the heart. The HMGB1 binds to the A/T-rich/MEF2-like sites of the cTnT promoter, as determined by chromatin immunoprecipitation assays. Mice carrying the −249bp promoter with point mutations disrupting the direct repeats expressed transgene at lower levels in the heart and ectopically in the brain. Ectopic expression of transgene was also observed in developing limbs and head. These results suggest an important role for the direct repeat in determining the cardiac specificity. Furthermore, mice carrying a mutant promoter simultaneously disrupting the direct repeats and overlapping GATA site failed to express the transgene in any tissues tested. Therefore, the direct repeat and overlapping GATA site are critical for the expression level and cardiac specificity. The F module controls one level of cardiac specificity. For a uniform and high level of cardiac-specific expression, the upstream element (−497bp to −250bp) is further required, possibly through the D enhancer module and the combination of Nkx2.5 and GATA sites.
登录
查看更多内容
影响因子：
8.3
作者：
SIGMUND, CD
通讯作者：
SIGMUND, CD
影响因子：
20.1
作者：
Song, Lanying;Yan, Wensheng;Jiao, Kai
通讯作者：
Jiao, Kai
影响因子：
11.4
作者：
Durocher, D;Charron, F;Nemer, M
通讯作者：
Nemer, M
影响因子：
10.5
作者：
Jiao, K;Kulessa, H;Hogan, BLM
通讯作者：
Hogan, BLM
影响因子：
5.3
作者：
Lee, Y;Shioi, T;Izumo, S
通讯作者：
Izumo, S