CREB-1alpha is recruited to and mediates upregulation of the cytochrome c promoter during enhanced mitochondrial biogenesis accompanying skeletal muscle differentiation.
To further understand pathways coordinating the expression of nuclear genes encoding mitochondrial proteins, we studied mitochondrial biogenesis during differentiation of myoblasts to myotubes. This energy-demanding process was accompanied by a fivefold increase of ATP turnover, covered by an eightfold increase of mitochondrial activity. While no change in mitochondrial DNA copy number was observed, mRNAs as well as proteins for nucleus-encoded cytochrome c, cytochrome c oxidase subunit IV, and mitochondrial transcription factor A (TFAM) increased, together with total cellular RNA and protein levels. Detailed analysis of the cytochrome c promoter by luciferase reporter, binding affinity, and electrophoretic mobility shift assays as well as mutagenesis studies revealed a critical role for cyclic AMP responsive element binding protein 1 (CREB-1) for promoter activation. Expression of two CREB-1 isoforms was observed by using specific antibodies and quantitative reverse transcription-PCR, and a shift from phosphorylated CREB-1Delta in myoblasts to phosphorylated CREB-1alpha protein in myotubes was shown, while mRNA ratios remained unchanged. Chromatin immunoprecipitation assays confirmed preferential binding of CREB-1alpha in situ to the cytochrome c promoter in myotubes. Overexpression of constitutively active and dominant-negative forms supported the key role of CREB-1 in regulating the expression of genes encoding mitochondrial proteins during myogenesis and probably also in other situations of enhanced mitochondrial biogenesis.
Franko
Andras
A
Institute of Vegetative Physiology, Medical Faculty, University of Köln, Robert-Koch-Strasse 39, 50931 Köln, Germany.
Mayer
Sabine
S
Thiel
Gerald
G
Mercy
Ludovic
L
Arnould
Thierry
T
Hornig-Do
Hue-Tran
HT
Wiesner
Rudolf J
RJ
Goffart
Steffi
S
D051697:Activating Transcription Factor 1 / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured / Q000378:metabolism; D017362:Cyclic AMP Response Element-Binding Protein / Q000502:physiology*; D045304:Cytochromes c / Q000096:biosynthesis / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics; D003576:Electron Transport Complex IV / Q000235:genetics; D018507:Gene Expression Regulation, Developmental / Q000235:genetics*; D017930:Genes, Reporter; D006801:Humans; D051379:Mice; D008931:Mitochondria, Muscle / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000096:biosynthesis / Q000235:genetics; D024510:Muscle Development / Q000235:genetics / Q000502:physiology*; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D032446:Myoblasts / Q000378:metabolism*; D010101:Oxygen Consumption; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D020033:Protein Isoforms / Q000502:physiology; D011499:Protein Processing, Post-Translational; D051381:Rats; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics; D015854:Up-Regulation
D051697:Activating Transcription Factor 1; C497100:Atf1 protein, mouse; C492004:Creb1 protein, mouse; D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; D020033:Protein Isoforms; C499631:TFAM protein, human; D014157:Transcription Factors; D045304:Cytochromes c; D003576:Electron Transport Complex IV