Evolutionary aspects of a new MyoD gene in amphioxus (Branchiostoma belcheri) and its promoter specificity in skeletal and cardiac muscles

Abstract

Vertebrate MyoD family of transcription factors contains four members including MyoD, Myf5, Myogenin and MRF4. These myogenic regulatory factors (MRFs) play key roles in regulating skeletal muscle development and growth. Evolutionary analysis suggests that the four vertebrate MRF genes were derived by gene duplications from a single ancestral gene during chordate evolution. Better understanding of the structure and regulation of MyoD expression in amphioxus Branchiostoma belcheri may provide insight into the evolutionary history of myogenic gene duplications because of the unique position of amphioxus in evolution. We report here that isolation and characterization of a new MyoD gene, AmphiMyoD, in B. belcheri. Sequence analysis revealed that the AmphiMyoD is more closely related to myogenic transcription factors in invertebrates and vertebrates compared with the previously identified three MyoD like genes in amphioxus, suggesting that the AmphiMyoD might be the closest relative of the ancestral myogenic gene. To determine if the AmphiMyoD gene promoter controls muscle-specific expression, the AmphiMyoD promoter was linked with the green fluorescence protein (GFP) reporter and the construct was microinjected into zebrafish embryos for transient expression assay. AmphiMyoD promoter directed skeletal muscle-specific GFP expression in zebrafish embryos. In addition, it also drove GFP expression in cardiac muscles of the injected embryos, but not in other non-muscle tissues. These data demonstrated that the AmphiMyoD promoter contained regulatory elements for skeletal and cardiac muscle-specific expression. Moreover, the regulatory element(s) could function across species.