Exon

About: Exon is a research topic. Over the lifetime, 38308 publications have been published within this topic receiving 1745408 citations. The topic is also known as: exons.

Expression and regulation of immunoglobulin heavy chain gene transfected into lymphoid cells.

Abstract: A plasmid including a mouse immunoglobulin mu gene was transfected into the IgG-secreting human lymphoid line HMy2 and mouse B- and pre-B-cell lines WEHI 231 and 18-81; stably transfected cells were selected. Transfected HMy2 cells synthesized mouse immunoglobulin mu chains as a major secreted protein but the WEHI 231 and 18-81 transfectants transcribed the introduced mu gene at lower levels. In HMy2 transfectants, most of the transcription of the introduced heavy chain gene initiated 40 and 62 bp upstream of the beginning of the VH exon translation start, although a small proportion of transcripts initiating further upstream was detected. WEHI 231 and 18-81 transfectants gave a much higher proportion of upstream initiation. Transient expression of the VH exon was monitored following transfection of mouse myeloma with the VH gene DNA in various plasmid constructs. VH transcription was only observed if the plasmids contained a segment derived from the large VH-CH intron of the immunoglobulin heavy chain locus. This segment, located between JH and switch regions, functioned both downstream of the VH exon and upstream in either orientation. The existence of a transcription enhancer element in this region is therefore proposed.

Coronaviruses Lacking Exoribonuclease Activity Are Susceptible to Lethal Mutagenesis: Evidence for Proofreading and Potential Therapeutics

Abstract: No therapeutics or vaccines currently exist for human coronaviruses (HCoVs). The Severe Acute Respiratory Syndrome-associated coronavirus (SARS-CoV) epidemic in 2002–2003, and the recent emergence of Middle East Respiratory Syndrome coronavirus (MERS-CoV) in April 2012, emphasize the high probability of future zoonotic HCoV emergence causing severe and lethal human disease. Additionally, the resistance of SARS-CoV to ribavirin (RBV) demonstrates the need to define new targets for inhibition of CoV replication. CoVs express a 3′-to-5′ exoribonuclease in nonstructural protein 14 (nsp14-ExoN) that is required for high-fidelity replication and is conserved across the CoV family. All genetic and biochemical data support the hypothesis that nsp14-ExoN has an RNA proofreading function. Thus, we hypothesized that ExoN is responsible for CoV resistance to RNA mutagens. We demonstrate that while wild-type (ExoN+) CoVs were resistant to RBV and 5-fluorouracil (5-FU), CoVs lacking ExoN activity (ExoN−) were up to 300-fold more sensitive. While the primary antiviral activity of RBV against CoVs was not mutagenesis, ExoN− CoVs treated with 5-FU demonstrated both enhanced sensitivity during multi-cycle replication, as well as decreased specific infectivity, consistent with 5-FU functioning as a mutagen. Comparison of full-genome next-generation sequencing of 5-FU treated SARS-CoV populations revealed a 16-fold increase in the number of mutations within the ExoN− population as compared to ExoN+. Ninety percent of these mutations represented A:G and U:C transitions, consistent with 5-FU incorporation during RNA synthesis. Together our results constitute direct evidence that CoV ExoN activity provides a critical proofreading function during virus replication. Furthermore, these studies identify ExoN as the first viral protein distinct from the RdRp that determines the sensitivity of RNA viruses to mutagens. Finally, our results show the importance of ExoN as a target for inhibition, and suggest that small-molecule inhibitors of ExoN activity could be potential pan-CoV therapeutics in combination with RBV or RNA mutagens.

High frequencies in African and non-African populations of independent mutations in the mannose binding protein gene

Abstract: We have previously identified, in three British families having an index child with frequent infections, a point mutation (GGC-->GAC) in codon 54 of exon 1 of the gene for the human lectin mannose binding protein (MBP). This was associated with low serum levels of this complement activating protein and would be anticipated to impair opsonization of mannose rich microorganisms. We now report a second point mutation (GGA-->GAA) in Gambians from West Africa, involving codon 57 of exon 1. By substituting carboxylic acids for axial glycines in the translated proteins both mutations would be expected to disrupt the secondary structure of the collagenous triple helix of the 96 kDa MBP subunits. In the Gambians the codon 57 mutation was studied by PCR, sequence analysis and restriction analysis and found to be remarkably common (frequency of the mutant gene 0.29 in adults and 0.23 in newborns) whereas the codon 54 mutation was very rare (frequency 0.003). However, the codon 54 mutation was frequent in both a British Caucasian and a Hong Kong Chinese population (frequency of the mutant gene 0.17 and 0.11 respectively). It was predicted that both homozygous and heterozygous individuals would have profoundly reduced serum levels of the protein and this was confirmed by immunoassay as was the reduced capacity of such sera to activate complement through the MBP initiated classical pathway. Our data indicate that the two mutations have arisen independently since the divergence of African and non African populations and both have attained high frequencies.