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D Kravis

Publications and source records attributed to D Kravis.

5 recordsLinked to original sources
PubMed1985

Quantitation of type II procollagen mRNA levels during chick limb cartilage differentiation.

A single-stranded DNA probe complementary to chicken type II procollagen mRNA has been used to quantitate levels of that mRNA present in chicken limb mesenchyme during cartilage differentiation. Excess labeled probe prepared from a cDNA template cloned in M13mp9 was hybridized to completion to increasing amounts of total RNA and assayed by protection from S1 nuclease digestion. Estimates of the absolute levels of type II procollagen RNA were determined using the M13mp9 template containing the coding strand as a standard. RNA complementary to the probe increased from 20 copies per diploid genome in stage 24 limb to approximately 2000 copies per diploid genome in stage 24 limb mesenchyme which had differentiated to cartilage in culture. Similar levels were found in cartilage from stage 31 limb. Sternal cartilage from 17-day embryos contained approximately 10,000 copies per diploid genome suggesting that the level of expression of this gene is different in limb growth cartilage compared with sternal cartilage. Low but detectable levels of RNA complementary to the probe were observed in limb at stages 20-24. Since a large fraction of the type II procollagen RNA in these early limbs is associated with polysomes, the type II procollagen gene appears to be expressed at a low level prior to phenotypic differentiation and prior to the accumulation of immunologically detectable levels of type II collagen.

Animals↗
PubMed1984

Structure and sequence of the chicken type II procollagen gene. Characterization of the region encoding the carboxyl-terminal telopeptide and propeptide.

The DNA sequence of two overlapping cDNA clones and a genomic lambda clone covering the region coding for 288 amino acids at the COOH terminus of the chicken type II procollagen gene is reported. This region consists of 4 exons coding for the last 15 amino acids of the triple helical domain and 273 amino acids which correspond to the COOH-terminal telopeptide and COOH-terminal propeptide. The sequence, base composition, and codon usage of this region of the type II procollagen gene show particularly high similarity to those of the chicken alpha 1(I) procollagen gene and differ from those of the alpha 2(I) and alpha 1(III) gene sequences. Two DNA tracts of low sequence similarity were observed. One of these regions spans the telopeptide and COOH-terminal propeptidase cleavage site, although 4-5 amino acids at the actual cleavage site are conserved compared with the alpha 1(I) and alpha 2(I) genes. A region of unusually high nucleotide sequence conservation is present in exon 2 (amino acids 171c - 186c ) consisting of approximately 45 nucleotides with only one or two base substitutions compared with the other procollagen genes. Within this conserved sequence is a site for carbohydrate attachment. The 3' nontranslated sequence of the type II procollagen mRNA is longer than that of either the alpha 1(I) or alpha 2(I) mRNA and contains several unusual long tracts consisting primarily of one or two bases. Although the canonical site for polyadenylation is not present, two related sequences, AACAAA and ATATAAA , are present 32 and 41 bases preceding the end of the major RNA species. The exon/intron structure of the type II procollagen gene is similar to that of other collagen genes which have been described. This DNA sequence provides the first extensive report of the amino acid sequence of chicken type II procollagen.

Amino Acid Sequence↗
PubMed1982

Construction and partial characterization of two recombinant cDNA clones for procollagen from chicken cartilage.

Type II procollagen mRNA has been partially purified from embryonic chick sternal cartilage by guanidine hydrochloride extraction, sucrose gradient sedimentation and Sepharose 4B chromatography. Double stranded cDNA was synthesized using AMV reverse transcriptase and E. coli DNA polymerase I, tailed using terminal transferase and inserted into the Pst I site of pBR322. Two putative type II procollagen cDNA clones have been characterized. Both plasmids hybridize to 2 sternal RNA species, a major species of 5.3 kb and a minor species of 7 kb. These RNAs are present in total RNA from sterna and differentiated limb bud cultures but are not detected in RNA from stage 24 limb bud which has not yet differentiated to cartilage or in RNA from calvaria. The time of appearance of these RNAs during the differentiation of limb mesenchyme in culture parallels the appearance of translatable type II procollagen mRNA.

Animals↗