B-Myb represses trans-activation of the Col5A2 collagen promoter indirectly via inhibition of binding of factors interacting with positive elements within the first exon.
B-myb, a member of the myb gene family, was originally isolated based on its high homology with c-myb in the DNA-binding domain. Previously we showed that B-myb is expressed in bovine vascular smooth muscle cells (SMCs) in a cell cycle-dependent fashion, and inhibits type I collagen gene promoter activity. Here, we have explored its role in regulation of another fibrillar collagen gene, Col5A2, encoding the (alpha2 chain of type V collagen. Ectopic expression of B-Myb decreased alpha 2(V) promoter activity and endogenous alpha 2(V) collagen mRNA levels. The responsive region of the alpha 2(V) collagen gene was localized to a fragment including 100 bp of basal promoter and 150 bp of exon 1 sequences, which contained two CRE-like elements. Binding to these elements increased upon deprivation of serum-growth factors, when expression of the Col5A2 gene is elevated, leading us to test their role despite the failure of excess unlabelled CRE oligonucleotide from the somatostatin gene to successfully compete for binding. Mutation of the elements significantly decreased the basal level of alpha2(V) collagen promoter activity and ablated inhibition by B-Myb. Furthermore, addition of B-Myb-glutathionine S-transferase fusion protein inhibited complex formation. Thus, these results confirm a major role for B-Myb in mediating intracellular signals controlling collagen gene expression in vascular SMCs. A model of indirect repression of the Col5A2 gene by B-Myb, via interaction with a positively-acting matrix regulatory factor, termed MRF-V, is discussed.
Kypreos
K E
KE
Department of Biochemistry, Boston University School of Medicine, MA 02118, USA.
Marhamati
D J
DJ
Sonenshein
G E
GE
D000818:Animals; D002417:Cattle; D018797:Cell Cycle Proteins; D002457:Cell Extracts; D002467:Cell Nucleus; D002478:Cells, Cultured; D002874:Chromosome Mapping; D003094:Collagen / Q000235:genetics*; D017124:Conserved Sequence; D016895:Culture Media, Serum-Free; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005091:Exons; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D009131:Muscle, Smooth, Vascular / Q000166:cytology; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
D018797:Cell Cycle Proteins; D002457:Cell Extracts; D016895:Culture Media, Serum-Free; D004268:DNA-Binding Proteins; C064352:MYBL2 protein, human; D012097:Repressor Proteins; D015534:Trans-Activators; D003094:Collagen