Shimizu N, Smith G, Izumo S
Department of Medicine, Harvard Medical School, Boston, MA 02215.
Nucleic Acids Res. 1993 Aug 25;21(17):4103-10. doi: 10.1093/nar/21.17.4103.
The A element, a fourteen base pair sequence in the rabbit myosin heavy chain (HC) beta promoter (-276/-263), contains the M-CAT motif, a cis-acting element found in several muscle-specific genes. The A element is essential for muscle-specific transcription of the myosin HC beta gene. Recently, we have identified both muscle-specific and ubiquitous factors (A1 and A2 factors, respectively) that bind to the A element. Since the sequence of the A element is very similar to the GTIIC motif in the SV40 enhancer, we examined the relationship between A-element-binding factors and a GTIIC binding factor TEF-1, recently isolated from HeLa cells. The GTIIC motif was bound by the A1 and A2 factors in muscle nuclear extracts and competed with the A element for DNA-protein complex formation. Antibody against human TEF-1 'supershifted' the ubiquitous A2 factor-DNA complex, but did not alter the mobility of the muscle-specific A1 factor-DNA complex. We isolated a murine cDNA clone (mTEF-1) from a cardiac cDNA library. The clone is highly homologous to Hela cell TEF-1. The in vitro transcription/translation product of mTEF-1 cDNA bound to the A element, and the DNA binding property of mTEF-1 was identical to that of the A2 factor. Transfection of mTEF-1 cDNA into muscle and non-muscle cells confirmed that mTEF-1 corresponds to A2, but not to A1 factors. The mTEF-1 mRNA is expressed abundantly in skeletal and cardiac muscles, kidney and lung, but it is also expressed at lower levels in other tissues. These results suggest that the M-CAT binding factors consist of two different factors; the ubiquitous A2 is encoded by mTEF-1, but the muscle-specific A1 factor is distinct from mTEF-1.
A元件是兔肌球蛋白重链(HC)β启动子(-276 / -263)中的一个14个碱基对的序列,包含M-CAT基序,这是一种在多个肌肉特异性基因中发现的顺式作用元件。A元件对于肌球蛋白HCβ基因的肌肉特异性转录至关重要。最近,我们鉴定了与A元件结合的肌肉特异性和普遍存在的因子(分别为A1和A2因子)。由于A元件的序列与SV40增强子中的GTIIC基序非常相似,我们研究了A元件结合因子与最近从HeLa细胞中分离出的GTIIC结合因子TEF-1之间的关系。肌肉核提取物中的A1和A2因子与GTIIC基序结合,并与A元件竞争DNA-蛋白质复合物的形成。抗人TEF-1抗体使普遍存在的A2因子-DNA复合物“超迁移”,但没有改变肌肉特异性A1因子-DNA复合物的迁移率。我们从心脏cDNA文库中分离出一个小鼠cDNA克隆(mTEF-1)。该克隆与HeLa细胞TEF-1高度同源。mTEF-1 cDNA的体外转录/翻译产物与A元件结合,mTEF-1的DNA结合特性与A2因子相同。将mTEF-1 cDNA转染到肌肉和非肌肉细胞中证实,mTEF-1对应于A2因子,而不是A1因子。mTEF-1 mRNA在骨骼肌、心肌、肾脏和肺中大量表达,但在其他组织中也有较低水平的表达。这些结果表明,M-CAT结合因子由两种不同的因子组成;普遍存在的A2由mTEF-1编码,但肌肉特异性A1因子与mTEF-1不同。