Analysis of cDNA and genomic clones coding for the proα1 chain of calf Type II collagen (original) (raw)
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Proceedings of the National Academy of Sciences, 1980
A series of overlapping recombinant clones, which cover the a2 (type I) collagen gene, have been isolated by stepwise screening of two libraries of chicken genomic DNA fragments. The first genomic clone was isolated by using a cloned cDNA containing a2 collagen DNA sequences as hybridization probe. The other clones were obtained by a sequence of screenings using defined fragments of the successive genomic clones as hybridization probes. Several types of experiments indicated that the DNA of these clones are truly overlapping and span 55 kilobase pairs of contiguous DNA sequences in the chicken genome. Sequence analysis of small DNA segments of some of these clones confirm that they contain coding sequences which specify a2 collagen. Electron microscopic analysis of hybrids between type I c2 collagen mRNA and the overlapping genomic clones indicates that the chicken c2 collagen gene has a length of at least 37 kilobases, about 7.4 times longer than the corresponding translatable cytoplasmic mRNA. The coding information for a2 collagen is distributed in more than 50 coding sequences which are interrupted by intervening sequences of various sizes. The structure of the gene implies that the conversion of precursor RNA to mature mRNA for cr2 collagen includes at least 50 splicing events.
Isolation and characterisation of a genomic clone encoding chick α1 type III collagen
Journal of Biological Chemistry
We have isolated a cloned segment of the chick alpha 1 type III collagen gene from a genomic DNA library. This DNA clone was obtained by cross-hybridization with a cDNA clone containing sequences coding for a segment of the carboxy propeptide of chick alpha 1 type I collagen. This genomic clone was identified as coding for alpha 1 type III collagen by DNA sequence analysis of two segments of the gene and by its ability to hybridize to crop mRNA but not to calvaria mRNA in agreement with the presence of alpha 1 type III collagen in crop but not in calvaria. The deduced amino acid sequences reveal characteristic features for alpha 1 type III collagen and are in good agreement with the known protein sequences of calf and human alpha 1 type III collagen. The size of alpha 1 type III mRNA is about 6 kilobases, slightly larger than the mRNA for alpha 1 type I. R-loop electron microscopic analysis indicates that at least 16 exons are interspersed in a 9-kilobase DNA segment that contains t...
Fine structural analysis of the chicken pro alpha 2 collagen gene
Proceedings of the National Academy of Sciences, 1981
Forty-two kilobase pairs of cloned chicken DNA containing 80% of the pro a2 (type I) collagen gene and 8 kilobase pairs of 3' flanking sequences have been isolated. Detailed analysis of these clones indicates that this collagen gene spans approximately 40 kilobase pairs of DNA and contains on te order of 50 introns. The fine structure of 40% of the pro a2 gene, including its 3' end, was determined by Southern blot restriction endonuclease mapping using a 2.6-kilobase pair procollagen cDNA clone, pCg45, as a probe, and by DNA sequence determination of more than 2 kilobase pairs of this part of the genome. Exons in the triple-helical coding region are all multiples of the 9 base pairs coding for the Gly-X-Y triplet and vary in size from 45 to 108 base pairs. The sequences of all six exons in a 3.8-kilobase pair EcoRI fragment were determined. One of these, a 249-base pair exon, joins the collagen domains; it codes for the last 15 amino acids of the triple-helica coding region, the telopeptide, and the first 53 amino acids of the carboxy-terminal propeptide. * To whom requests for details of DNA sequence analysis should be addressed. t To whom reprint requests should be addressed. 712
The Journal of biological chemistry, 1991
A series of cDNA clones corresponding to the 5' end of the chicken type II collagen mRNA were generated using a single-sided polymerase chain reaction technique. Analysis of these cDNAs showed that the second exon of the gene is alternatively spliced such that it is either present or absent in the mRNA. This exon encodes a 70-amino acid cysteine-rich globular domain which is present in the amino-terminal propeptides of alpha 1(I), alpha 1(III), and alpha 2(V) procollagen chains but which was previously thought to be absent from type II procollagen. Analysis of the expression of the two alternatively spliced forms of the chicken type II collagen mRNA showed that the mRNA without the second exon was the predominant form (approximately 90%) in sternal cartilage from 14-day embryos, but in precartilage limb mesenchyme only the form including the second exon was detected. This later form was also present in a number of non-cartilage tissues including embryonic calvaria, skin, heart, ...
Analysis of the promoter region and the N-propeptide domain of the human proα2(I) collagen gene
Nucleic Acids Research, 1985
We have located the exon coding for the start site of transcription of the human proa2(I) collagen gene. Comparison with the homologous region of other fibrillar collagen genes has confirmed the existence of a consensus sequence (CATGTCTA-n-TAGACATG) capable of forming a hairpin secondary structure possibly involved in the regulation of collagen biosynthesis. Sequence comparison of the chromosomal regions at the 5' end of the proal(I) and proa2(I) collagen genes failed to identify unique DNA elements potentially mediating common regulatory signals. Sequencing of four exons coding for the Nterminal propeptide has determined most of its structure and it has implied the existence of smaller coding units similar to the 11 and 18 bp exons originally described in the avian gene.
Gene, 1988
Dysfunctions of the genes coding for the two chains of the human type-1 procollagen result in genetic disorders that affect the integrity of bone, ligaments, tendons, and other connective tissues. While the primary amino acid (aa) sequence of one of the two type-1 subunits, proa2(1), has been derived in its entirety from the analysis of overlapping cDNAs, the sequence of the first 247 aa residues of the helical domain of the other polypeptide, proal(had yet to be determined. To this end, we have sequenced nearly 4 kb of the human proal(1) collagen gene and identified twelve open reading frames whose conceptual amino acid translation exhibits 95 yO homology to the first 247 aa of rat CI l(1) chain. Furthermore with these and other data, some of which previously unpublished, we have derived the complete sequence of the first 7618 bp of the gene. This region comprises the 25 exons encoding the N-terminal pre-propeptide and five of the eight cyanogen-bromidederived peptides. This information therefore represents a most useful reference for the characterization of molecular defects in individuals affected by various connective tissue disorders.
Accurate in vitro transcriptional initiation of the chick alpha 2 (Type I) collagen gene
The Journal of biological chemistry, 1981
Chick genomic DNA containing the extreme 5' end of the alpha 2 (type I) collagen gene has been used as template in an in vitro HeLa cell transcription system. RNA polymerase II-dependent transcription initiates from a specific site on this DNA. The precise location of this site was determined by three types of experiments: sizing of in vitro-synthesized RNA runoff transcripts, comparing the sequence of the in vitro-made RNA transcripts with the structure of the DNA template, and identifying the first and second nucleotides of the in vitro-synthesized transcripts. Transcription was found to initiate 33 base pairs downstream from a canonical Goldberg-Hogness sequence (TATAAATA). This in vitro start site is the same as the initiation site of in vivo-synthesized collagen RNA.
Biochemistry, 1977
The covalent structure of the first 111 residues from the N-terminus of peptide al(II)-CB10 from bovine nasal-cartilage collagen is presented. This region comprises residues 552-661 of the al(II) chain. The sequence was determined by automated Edman degradation of peptide al(II)-CB10 and of peptides produced by cleavage with trypsin and hydroxylamine. Comparison of this region of the al(II) chain with the homologous segment of the al(I) chain indicated a homology level of 85 %, slightly higher than that of 81 % reported for the N-terminal region of the al(II) chain (Butler, Miller & Finch (1976) Biochemistry 15, 3000-3006). The occurrence of two residues of glycosylated