RESUMEN
We have isolated overlapping clones containing the 5'-terminal portion of the human pro-alpha 1(III) collagen gene (COL3A1). This has enabled us to extend our previous studies and thus generate a restriction map of nearly 64 kb of DNA encompassing all of COL3A1 and more than 20 kb of flanking sequences. Aside from the complete nucleotide and amino acid sequences of type-III N-pre-propeptide, this study has established the number of the corresponding exons, whose relative organization deviates from the pattern observed in the analogous regions of type-I procollagen genes, COL1A1 and COL1A2. Moreover, we have sequenced 1628 bp of the 5'-flanking region of COL3A1, from the transcription start point (tsp) to an AluI repetitive element. Pairwise comparison with the analogous segment of the mouse gene has showed 78% sequence similarity in nearly 270 bp immediately preceding the tsp and including the TATA element and a presumptive NF-1 binding site. Relatively close to the tsp, but upstream from the region of homology with the murine gene, a potential AP-1 binding site has also been identified.
Asunto(s)
ADN/genética , Genes , Procolágeno/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Evolución Biológica , Clonación Molecular , Procesamiento Automatizado de Datos , Exones , Humanos , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Fragmentos de Péptidos/genética , Plásmidos , Regiones Promotoras Genéticas , Mapeo Restrictivo , Homología de Secuencia de Ácido NucleicoRESUMEN
The organization of the exons coding for the N-terminal portion of human type II procollagen has been determined. Aside from inferring the previously unknown primary structure of type II N-propeptide, this study has revealed that this coding domain of the gene exhibits an organization uniquely distinct from those of type I and type III collagens. This finding substantiates the notion that the N-propeptide coding domains of the fibrillar collagen genes evolved under less stringent selection than those encoding the C-propeptide and triple helical regions.
Asunto(s)
Procolágeno/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Evolución Biológica , Bovinos/genética , Colágeno/genética , Exones , Genes , Humanos , Datos de Secuencia Molecular , Homología de Secuencia de Ácido Nucleico , Especificidad de la EspecieRESUMEN
We have determined the nucleotide sequence of a cDNA clone encoding the amino-terminal portion of human alpha 2(V) procollagen and found that the structure of the 186-residue amino-terminal propeptide closely resembles those of the fibril-forming procollagens. Juxtaposed to a 26-residue leader peptide, pro-alpha 2(V) exhibits a characteristic cysteine-rich globular region followed by 24 Gly-X-Y repeats which are interrupted by two short non-collagenous sequences. Upon closer examination, each of these two sequences was noted to display structural motifs characteristic of either pro-alpha 1(I) and pro-alpha 1(III) collagens or pro-alpha 1(II) collagen, respectively. Finally, within the amino-terminal telopeptide, a putative amino-terminal proteinase cleavage site, Ala-Gln, was identified. This latter finding strongly suggests that the alpha 2(V) amino-terminal propeptide can be potentially processed and thus leaves unresolved the issue pertaining to the nature of the collagenase-resistant sequence that is retained by mature type V collagen molecules.
Asunto(s)
Procolágeno/genética , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , ADN/genética , ADN/aislamiento & purificación , Femenino , Genes , Humanos , Datos de Secuencia Molecular , Placenta/metabolismo , Embarazo , Mapeo Restrictivo , Homología de Secuencia de Ácido NucleicoRESUMEN
Sixty kilobases of cloned DNA containing the entire human pro-alpha 2(I) collagen gene and 22 kilobases of flanking sequences have been isolated. Like the homologous avian gene, the 1366 amino acid residues of the human pre-pro-alpha 2(I) chain are encoded by 52 exons, whose relative locations and sizes have been determined. Analysis of the 5'- and 3'-untranslated regions have confirmed their exact lengths, as well as conclusively established the nature of five polymorphic mRNA transcripts.
Asunto(s)
Genes , Procolágeno/genética , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , ADN/metabolismo , Enzimas de Restricción del ADN , Humanos , Datos de Secuencia Molecular , Peso Molecular , Conformación de Ácido Nucleico , Mapeo NucleótidoRESUMEN
A bovine cDNA library constructed from fetal cartilage RNA was screened with a pro alpha 1(II) collagen specific chicken cDNA. A recombinant clone (Bc 7), with an insert of 1 kb, was identified and shown to contain sequences exhibiting 85% homology with the chicken pro alpha 1(II) collagen C-propeptide. Interspecies comparison strongly suggested that one potential glycosylation site present in the avian C-propeptide is not utilized, since this site is absent in the bovine chain. In addition, two overlapping genomic clones (Pal 3 and Pal 4) were isolated and partially characterized. These clones span 23 kb of DNA and contain approximately 17 kb of the pro alpha 1(II) calf gene. Sequencing of exon 1 has determined the length of the 3' untranslated region and the exact location of the polyadenylation attachment site.
Asunto(s)
Colágeno/genética , ADN/análisis , Secuencia de Aminoácidos , Animales , Cartílago/análisis , Bovinos , Pollos , Clonación Molecular , Enzimas de Restricción del ADN/metabolismo , Hibridación de Ácido Nucleico , Poli A/metabolismo , ARN/metabolismo , ARN MensajeroRESUMEN
Using a cDNA probe specific for the bovine Type II procollagen, a series of overlapping genomic clones containing 45 kb of contiguous human DNA have been isolated. Sequencing of a 54 bp exon, number 29, provided direct evidence that the recombinant clones bear human Type II collagen sequences. Localization of the 5' and 3' ends of the gene indicated that the human Type II collagen gene is 30 kb in size. This value is significantly higher than that of the homologous avian gene. The segregation of a polymorphic restriction site in informative families conclusively demonstrated that the Type II gene is found in a single copy in the human haploid genome. Finally, sequencing of a triple helical domain exon has confirmed that a rearrangement leading to the fusion of two exons occurred in the pro alpha 1(I) gene, following the divergence of the fibrillar collagens.