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Expression of the fibroblast growth factor receptor 1 (FGFR1) gene in skeletal muscle is positively regulated in proliferating myoblasts and declines during differentiation. We have characterized the cis-regulatory elements in the proximal region of the FGFR1 promoter which render positive transcriptional activity. Multiple elements between ؊69 and ؊14 activate the FGFR1 promoter. Myoblast transfections revealed that potential Sp transcription factor binding sites are required for promoter activity. Electromobility shift assays indicated that myoblast nuclear proteins specifically bind to these cis-elements and that differentiated myotube nuclear extracts do not form these same complexes. In addition, Southwestern blot analysis detected binding of the most proximal Sp motif to a Sp1-like protein present in myoblast nuclear extracts but not in myotubes. In corroboration, Sp1 and Sp3 proteins were detected only in myoblasts and not in differentiated myotubes. Finally, transfection of Drosophila SL2 cells showed that Sp1 is a positive regulator of FGFR1 promoter activity and that Sp3 is a coactivator via the proximal Sp binding sites. These studies demonstrate that the FGFR1 promoter is activated by Sp transcription factors in proliferating myoblasts and demonstrate at least part of the mechanism by which FGFR1 gene expression is downregulated in differentiated muscle fibers.During vertebrate myogenesis, mesodermally derived cells within myogenic lineages proliferate as mononucleated myoblasts before differentiation into postmitotic, multinucleated muscle fibers. Both the sustained proliferation of myoblasts and subsequent withdrawal from the cell cycle as a part of differentiation are regulated by signal transduction cascades initiated by environmental signals including growth factors. Members of the fibroblast growth factor (FGF) 1 family of signaling molecules are capable of sustaining myoblast proliferation and delaying differentiation. The cellular effects of FGF signaling are mediated through a small family of fibroblast growth factor receptors (FGFRs). FGF1 and FGF2 possess well documented mitogenic activity for skeletal myoblasts, and these factors bind to FGFR1 in the cell surface of proliferating myoblasts. In addition to skeletal muscle myoblasts, FGFR1 is also expressed during development of the brain, skin, bones, and cardiac muscle (1). It has recently been shown that FGFR1 can be translocated to the nucleus via importin B and that it plays an important role in the regulation of the cell cycle by inducing nuclear target genes (2). However, FGF signaling declines during myoblast differentiation. This decline is the result of loss of cell surface receptor and a coordinate decrease in FGFR1 mRNA (3-5).The significance of the developmentally regulated expression of FGFR1 in relation to muscle growth and patterning in vivo has been partially examined. Myoblast differentiation and muscle fiber formation were delayed in chick limb musculature overexpressing wild type FGFR1. Conversely, premature differentiat...

Developmentally controlled transcriptional regulation of myogenic cell proliferation and differentiation via expression of the fibroblast growth factor receptor 1 (FGFR1) gene is positively regulated by Sp1 and negatively regulated by E2F4-based transcriptional complexes. We report that p107 and p130 formed transcriptional complexes with E2F4 on the FGFR1 promoter and repressed FGFR1 gene transcription in myogenic cells. However, in Drosophila melanogaster SL2 cells, only p107 was able to repress Sp1-mediated transactivation of the FGFR1 promoter. Gel shift assays using transfected myoblast nuclear extracts showed that ectopic p107 reduced Sp1 occupancy of the proximal Sp binding site of the FGFR1 promoter, and coimmunoprecipitation studies indicated that Sp1 interacts with p107 but not with p130. Gel shift assays also demonstrated that Sp1 interacted with p107 in E2F4-p107 transcriptional complexes in myoblasts. The nature of the repressor transcriptional complex was altered in differentiated muscle fibers by the relative loss of the E2F4-p107-Sp1 transcription complex and replacement by the repressor E2F4-p130 complex. These findings demonstrate that activation and repression of FGFR1 gene transcription is governed by interplay between Sp1, p107, p130, and E2F4 in distinct transcriptional complexes during skeletal muscle development. Fibroblast growth factor receptors (FGFRs)1 have diverse functional roles in mitogenesis, angiogenesis, cell migration, differentiation, mesoderm induction, bone growth and limb development (1). In skeletal muscle, FGFR1 mediates the mitogenic activity initiated by FGF1 and FGF2. During skeletal myogenesis, FGFR1 gene expression is positively regulated in proliferating myoblasts and negatively regulated in differentiated muscle fibers. The functional significance of regulated expression of the FGFR1 gene during myogenesis is demonstrated by overexpression in vivo. Chick embryos overexpressing wild-type FGFR1 displayed delayed myoblast differentiation and muscle fiber formation. On the contrary, chick embryos overexpressing a dominant-negative form of FGFR1 displayed premature muscle fiber formation with decreased muscle mass (2, 3).Although regulation of FGFR1 gene expression is important for normal growth and development of skeletal muscles, the molecular mechanism governing its transcription is poorly understood. Positive regulation of FGFR1 gene expression in proliferating myoblasts is governed by the Sp1 transcription factor. The chicken FGFR1 promoter contains two functional, distal Sp1 binding sites, and the proximal region contains three Sp1 binding sites, all of which are essential for full promoter activity in proliferating myoblasts (4, 5). Negative transcriptional regulators control FGFR1 promoter activity as FGFR1 gene expression declines during myogenic differentiation. We recently identified E2F4 as a negative regulator of FGFR1 gene expression in skeletal muscle cells (6). Its repressor activity was mediated by E2F4 binding to a proximal cis-element at Ϫ65 bp. Howev...

Background: FGFR1 gene expression regulates myoblast proliferation and differentiation, and its expression is controlled by Krüppel-like transcription factors. Results: KLF10 interacts with the FGFR1 promoter, repressing its activity and cell proliferation. Conclusion: KLF10 represses FGFR1 promoter activity and thereby myoblast proliferation. Significance: A model of transcriptional control of chicken FGFR1 gene regulation during myogenesis is presented.

Fibroblast growth factor receptor 1 (FGFR1) gene expression is positively and negatively regulated during muscle differentiation. We recently reported that FGFR1 gene expression was up-regulated by Sp transcription factors in proliferating myoblasts. However, the mechanism of down-regulation of this gene during differentiation is unknown. We have identified the transcription factor E2F4 as a negative regulator of FGFR1 gene expression. Immunodetection studies revealed that endogenous E2F1 and E2F2 proteins were cytoplasmic in myoblasts and myotubes, whereas E2F4 was abundant in the nuclei of both. Upon overexpression, E2F4 repressed FGFR1 promoter activity in a dose-dependent manner in myoblasts and Drosophila SL2 cells, and mutation of the E2F4 binding site increased FGFR1 promoter activity and reduced E2F4-mediated repression. Gel shift assays detected E2F4 binding to a synthetic

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