The entire primary structure of the murine alpha 1(VI) collagen chain was deduced from cloned cDNA. The predicted polypeptide consists of 1025 amino acids and shows extensive homology with the corresponding human and chicken chains. A genomic clone isolated with a cDNA probe was found to contain about 13 kilobases of the 5'-flanking region and the first and second exon, coding for the 5'-untranslated sequence and signal peptide and part of the N-terminal portion of the mature protein, respectively. Polymerase chain reaction and primer extension analyses revealed two major and several minor transcription start sites distributed over 76 base pairs (bp). The region just upstream of the transcription initiation sites lacks canonical TATA and CAAT boxes and Sp1 binding sites, but contains putative binding sites for other transcription factors and a 90-bp polypyrimidine tract with elements of dyad symmetry. Chimeric constructs were derived from different fragments of the 5'-flanking genomic region and the chloramphenicol acetyltransferase (CAT) gene and expression of the reporter gene was assayed following transfection of various cell types. A construct containing sequences extending from -215 to +41 directed high levels of CAT expression. The data indicate that this region harbours a functional promoter.

Murine alpha1(VI) collagen chain. Complete amino acid sequence and identification of the gene promoter region.

BONALDO, PAOLO;PICCOLO, STEFANO;VOLPIN, DINO;BRESSAN, GIORGIO
1993

Abstract

The entire primary structure of the murine alpha 1(VI) collagen chain was deduced from cloned cDNA. The predicted polypeptide consists of 1025 amino acids and shows extensive homology with the corresponding human and chicken chains. A genomic clone isolated with a cDNA probe was found to contain about 13 kilobases of the 5'-flanking region and the first and second exon, coding for the 5'-untranslated sequence and signal peptide and part of the N-terminal portion of the mature protein, respectively. Polymerase chain reaction and primer extension analyses revealed two major and several minor transcription start sites distributed over 76 base pairs (bp). The region just upstream of the transcription initiation sites lacks canonical TATA and CAAT boxes and Sp1 binding sites, but contains putative binding sites for other transcription factors and a 90-bp polypyrimidine tract with elements of dyad symmetry. Chimeric constructs were derived from different fragments of the 5'-flanking genomic region and the chloramphenicol acetyltransferase (CAT) gene and expression of the reporter gene was assayed following transfection of various cell types. A construct containing sequences extending from -215 to +41 directed high levels of CAT expression. The data indicate that this region harbours a functional promoter.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11577/123641
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