Complementary DNA

About: Complementary DNA is a research topic. Over the lifetime, 55301 publications have been published within this topic receiving 2752650 citations. The topic is also known as: cDNA & DNA, Complementary.

Molecular cloning of the beta-subunit of human prolyl 4-hydroxylase. This subunit and protein disulphide isomerase are products of the same gene.

Abstract: Prolyl 4-hydroxylase (EC 1.14.11.2), an alpha 2 beta 2 tetramer, catalyses the formation of 4-hydroxyproline in collagens by the hydroxylation of proline residues in peptide linkages. We report here the isolation of cDNA clones coding for the beta-subunit of prolyl 4-hydroxylase from a human hepatoma lambda gt11 library and a corresponding human placenta library. Five overlapping clones covering all the coding sequences and almost all the non-coding sequences were characterized. The size of the mRNA hybridizing with these clones in Northern blotting is approximately 2.5 kb. The clones encode a polypeptide of 508 amino acid residues, including a signal peptide of 17 amino acids. These human sequences were found to be very similar to those recently reported for rat protein disulphide isomerase (EC 5.3.4.1). The degree of homology between these two proteins was 84% at the level of nucleotide sequences or 94% at the level of amino acid sequences. Southern blot analyses of human genomic DNA with a cDNA probe for the beta-subunit indicated the presence of only one gene containing these sequences. The product of a single gene thus appears to possess two different enzymatic functions depending on whether it is present in cells in monomer form or in the prolyl 4-hydroxylase tetramer.

Complementation of a mutant cell line : central role of the 91 kDa polypeptide of ISGF3 in the interferon-alpha and -gamma signal transduction pathways

Abstract: Mutants in complementation group U3, completely defective in the response of all genes tested to interferons (IFNs) alpha and gamma, do not express the 91 and 84 kDa polypeptide components of interferon-stimulated gene factor 3 (ISGF3), a transcription factor known to play a primary role in the IFN-alpha response pathway. The 91 and 84 kDa polypeptides are products of a single gene. They result from differential splicing and differ only in a 38 amino acid extension at the C-terminus of the 91 kDa polypeptide. Complementation of U3 mutants with cDNA constructs expressing the 91 kDa product at levels comparable to those observed in induced wild-type cells completely restored the response to both IFN-alpha and -gamma and the ability to form ISGF3. Complementation with the 84 kDa component similarly restored the ability to form ISGF3 and, albeit to a lower level, the IFN-alpha response of all genes tested so far. It failed, however, to restore the IFN-gamma response of any gene analysed. The precise nature of the DNA motifs and combination of factors required for the transcriptional response of all genes inducible by IFN-alpha and -gamma remains to be established. The results presented here, however, emphasize the apparent general requirement of the 91 kDa polypeptide in the primary transcriptional response to both types of IFN.

Molecular cloning of complementary DNA encoding the lignin-forming peroxidase from tobacco: molecular analysis and tissue-specific expression

Abstract: Plant peroxidases play a major role in lignin formation and wound healing and are believed to be involved in auxin catabolism and defense to pathogen attack. The function of the anionic peroxidase isozymes is best understood in tobacco. These isozymes catalyze the formation of the lignin polymer and form rigid cross-links between lignin, cellulose, and extensin in the secondary plant cell wall. We report the purification of the anionic peroxidase isozymes from tobacco and their partial amino acid sequence. An oligonucleotide probe deduced from the amino acid sequence was used to screen a tobacco leaf cDNA library and a 1200-base-pair cDNA clone was isolated and sequenced in its entirety. The predicted amino acid sequence revealed a 22-amino acid signal peptide and a 302-amino acid mature protein (Mr, 32,311). The amino acid sequence was compared to that of the cationic peroxidases from horseradish and turnip and was found to be 52% and 46% homologous, respectively. By RNA blot analysis, the messenger for the tobacco isozyme was found to be abundant in stem tissue while expressed at very low levels in leaf and root tissue. Four distinguishable copies of the gene were found on genomic DNA blots. The gene copy number may reflect the allotetraploid nature of Nicotiana tabacum.