Latinkic Branko V, Cooper Brian, Smith Stuart, Kotecha Surendra, Towers Norma, Sparrow Duncan, Mohun Timothy J
Division of Developmental Biology, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, UK.
Development. 2004 Feb;131(3):669-79. doi: 10.1242/dev.00953. Epub 2004 Jan 7.
The mechanisms by which transcription factors, which are not themselves tissue restricted, establish cardiomyocyte-specific patterns of transcription in vivo are unknown. Nor do we understand how positional cues are integrated to provide regionally distinct domains of gene expression within the developing heart. We describe regulation of the Xenopus XMLC2 gene, which encodes a regulatory myosin light chain of the contractile apparatus in cardiac muscle. This gene is expressed from the onset of cardiac differentiation in the frog embryo and is expressed throughout all the myocardium, both before and after heart chamber formation. Using transgenesis in frog embryos, we have identified an 82 bp enhancer within the proximal promoter region of the gene that is necessary and sufficient for heart-specific expression of an XMLC2 transgene. This enhancer is composed of two GATA sites and a composite YY1/CArG-like site. We show that the low-affinity SRF site is essential for transgene expression and that cardiac-specific expression also requires the presence of at least one adjacent GATA site. The overlapping YY1 site within the enhancer appears to act primarily as a repressor of ectopic expression, although it may also have a positive role. Finally, we show that the frog MLC2 promoter drives pan myocardial expression of a transgene in mice, despite the more restricted patterns of expression of murine MLC2 genes. We speculate that a common regulatory mechanism may be responsible for pan-myocardial expression of XMLC2 in both the frog and mouse, modulation of which could have given rise to more restricted patterns of expression within the heart of higher vertebrates.
转录因子本身并非组织特异性的，但它们在体内建立心肌细胞特异性转录模式的机制尚不清楚。我们也不了解位置线索是如何整合的，以在发育中的心脏内提供区域不同的基因表达域。我们描述了非洲爪蟾XMLC2基因的调控，该基因编码心肌收缩装置的一种调节性肌球蛋白轻链。该基因在青蛙胚胎心脏分化开始时就开始表达，并且在心脏腔形成之前和之后的所有心肌中都有表达。通过在青蛙胚胎中进行转基因操作，我们在该基因的近端启动子区域内鉴定出一个82 bp的增强子，它对于XMLC2转基因的心脏特异性表达是必需且充分的。这个增强子由两个GATA位点和一个复合的YY1/CArG样位点组成。我们表明低亲和力的SRF位点对于转基因表达至关重要，并且心脏特异性表达还需要至少一个相邻的GATA位点的存在。增强子内重叠的YY1位点似乎主要作为异位表达的抑制因子起作用，尽管它也可能具有积极作用。最后，我们表明青蛙MLC2启动子驱动转基因在小鼠中的全心肌表达，尽管小鼠MLC2基因的表达模式更具局限性。我们推测一种共同的调控机制可能负责青蛙和小鼠中XMLC2的全心肌表达，对其进行调节可能导致高等脊椎动物心脏内更具局限性的表达模式。
