Biomedical subjects

B Coulombe

Publications and source records attributed to B Coulombe.

22 records · Page 2Linked to original sources

PubMed1988

Regulation of interferon gene expression: mechanism of action of the If-1 locus.

We have examined the mechanism of action of the If-1 interferon (IFN) regulatory locus. This locus controls the level of circulating IFN produced in inbred mice in response to intravenous injection of Newcastle disease virus. Mice carrying the If-1h (high) allele show circulating IFN levels 10- to 15-fold higher than those carrying the If-1l (low) allele. In this report we show that induced splenocytes from If-1h and If-1l mice produce IFN at levels which are in the same proportions as those found in the circulation. Higher levels of IFN-specific mRNA were observed in splenocyte populations from If-1h animals. This was due to increased transcription of IFN genes. At the same time, the high- and low-producing populations showed no significant difference in the number of IFN mRNA-containing cells. We conclude that the effect of If-1 in the spleen is to control the levels of transcription of the IFN genes in individual induced splenocytes.

Alleles↗

PubMed1988

Presence of transcription regulatory elements within an intron of the virus-inducible murine TIMP gene.

The expression of the gene for the murine tissue inhibitor of metalloproteinases (TIMP) is induced in response to viruses, growth factors, and phorbol esters. In this report we show that the accumulation of TIMP mRNA after Newcastle disease virus induction is caused by transcriptional activation of the gene. Comparison of the sequences of cDNA and genomic clones along with RNase protection and primer extension analyses revealed that the murine TIMP gene possesses multiple cap sites and that the exon 1 consists exclusively of 5'-noncoding sequences. We observed that DNA regions analogous to those found upstream of the virus-inducible interferon genes are present within intron 1 of the TIMP gene. To investigate the possible role of TIMP intron 1 in gene expression, we used a functional assay based on the transfection of plasmids in which the DNA segment to be tested is placed in proximity to a marker gene driven by the heterologous herpes simplex virus thymidine kinase promoter. Our results indicate that TIMP intron 1 contains DNA sequence elements capable of modulating the activity of a heterologous promoter in two different ways: (i) by enhancing constitutive expression and (ii) by conferring virus inducibility. These results suggest that intron 1 may be involved in the transcriptional regulation of TIMP gene expression.

Animals↗

PubMed1987

Characterization and expression of a murine gene homologous to human EPA/TIMP: a virus-induced gene in the mouse.

A genomic clone encompassing the entire coding region of a murine gene homologous to human erythroid potentiating activity/tissue inhibitor of metalloproteinase (EPA/TIMP) was isolated and sequenced. Based on alignment with human EPA/TIMP cDNAs we deduce a structure comprising five exons and four introns extending over 4.3 kb of DNA. In mouse and hamster cell lines transcription from this gene and interferon genes is induced by Newcastle Disease virus (NDV). Examination of the 5'-flanking sequences of the gene reveals a set of repeated elements with structural similarity to those previously described as inducer-responsive elements in the human IFN-beta 1 gene. The 4.3-kb DNA fragment encompassing the homologous murine EPA/TIMP gene was transfected into human T98G cells and transfectants tested for NDV inducibility. In contrast to the endogenous human gene, the integrated murine EPA/TIMP gene was NDV-inducible and TIMP activity was detectable in the cell culture fluid.

Amino Acid Sequence↗

PubMed1986

Expression of a synthetic human interferon-alpha 1 gene with modified nucleotide sequence in mammalian cells.

We have constructed a recombinant interferon (IFN) gene whose coding region is a chemically synthesized human (Hu)IFN-alpha 1 gene [Edge et al., Nature 292 (1981) 756-762] encoding the same protein as its natural counterpart while possessing important modifications. These modifications were introduced in order to: change codon usage, destroy integrity of repeated and complementary oligodeoxynucleotides in the coding region, introduce an intron, substitute other sequences for the IFN specific 3'- and 5'-flanking sequences and remove the leader sequence (which encodes for the signal peptide). We report here on the transfection and isolation of murine cells which contain low-copy-number insertions of our recombinant gene. These lines maintain a high steady-state level of biologically active HuIFN-alpha in the cytoplasm, produced from mRNAs of Mr expected for correctly processed transcripts. These results show directly that the leader sequence is essential for secretion of IFN proteins. They also argue against a crucial role of the particular features of the transcribed sequence of the natural HuIFN-alpha 1 gene in regulating its expression.

Animals↗