魚類肌肉調控蛋白MYF-5及MYOGENIN的生物特性：在胚胎早期發育中的角色及互動(2/2)

Abstract

Myf-5, a transcription factor that controls muscle differentiation, is expressed in somites during early embryogenesis. However, gene regulation of myf-5 is poorly understood and detailed functional analysis of the regulatory cis-elements is needed. In zebrafish, the myf-5 upstream sequence from -82 to -62 (-82/-62) was fused with a basal promoter and transferred to fertilized zebrafish eggs. The -82/-62 cassette drove GFP reporter gene expression specifically in the somites. Moreover, GFP signals were detected exclusively in the somites of 28-hpf embryos derived from eggs injected with 5×(-82/-62)/CMV/EGFP, which contained 5 copies of the -82/-62 cassette inserted within CMV promoter/enhancer. Thus, the -82/-62 cassette has dual functions: to enhance somite-specific expression and to repress non-specific expression during the early development of zebrafish embryos. This cassette sequence was highly conserved in mouse myf-5 and had the same function. Mutated sequence analysis and electrophoretic mobility shift assay of -82/-62 cassette showed that the -70/-62 sequence is the major element for controlling myf-5 specificity and for forming the DNA-protein complex. However, the putative CCAAT-like box, located at -66/-62, could not direct somite-specific expression. Therefore, the -70/-67 sequence is essential for controlling somite-specific expression and the CCAAT-like box is essential for activating gene transcription.