Cloning and characterization of the full length cDNA encoding alpha2 type I collagen of bullfrog Rana catesbeiana.
The present study determined nucleotide sequences of the full-length cDNA of alpha2 chain of bullfrog type I collagen. Hybridization of a bullfrog cDNA library with human alpha1 type I collagen cDNA yielded a clone named 6A-1 which was 3449 bp long and lacked a 5' region of the gene. A 5' region containing the translation initiation site was amplified by the reverse transcription polymerase chain reaction using poly(A)+RNA from tadpole tail tissues as template, and oligonucleotides encoding the translation initiation region of mammalian fibrillar collagens and the Gly-X-Y repeat region of clone 6A-1 as primers. As a result we obtained a 1518 bp long clone Y31. A 355 bp long clone Y31-9 was produced by extending clone Y31 from its ATG codon to a 127 bp upstream region. Combining these three clones, the complete nucleotide sequence of the full-length cDNA was determined which contained 4692 bp as a whole and 4065 bp in the open reading frame. The comparison of its structure with known collagen cDNAs of various vertebrates showed that the cDNA obtained codes for alpha2(I) chain of bullfrog. Its deduced amino acid sequence revealed the complete conservation of seven cysteine residues in the C-propeptide and three lysine residues in the N-telopeptide through the helical domain. Northern blot analysis revealed that the thyroid hormone regulated the expression of alpha2(I) collagen in an organ-dependent manner: intense up-regulation in the back skin and intestine, weak and transient up-regulation in the liver, and initial down-regulation, but later up-regulation in the tail. Prolactin increased its expression in both the back skin and tail. These results suggested that the expression of bullfrog alpha2(I) collagen is cooperatively regulated by these two metamorphosis-regulating hormones.
Asahina
K
K
Department of Biological Science, Faculty of Science, Hiroshima University, Japan.
Oofusa
K
K
Obara
M
M
Yoshizato
K
K
D000595:Amino Acid Sequence; D000818:Animals; D002645:Chickens; D003001:Cloning, Molecular; D003094:Collagen / Q000235:genetics*; D018076:DNA, Complementary; D005786:Gene Expression Regulation; D006801:Humans; D007448:Invertebrates; D008322:Mammals; D008675:Metamorphosis, Biological; D008969:Molecular Sequence Data; D011388:Prolactin / Q000502:physiology; D011892:Rana catesbeiana; D017386:Sequence Homology, Amino Acid; D013963:Thyroid Hormones / Q000502:physiology
D018076:DNA, Complementary; D013963:Thyroid Hormones; D011388:Prolactin; D003094:Collagen