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On the linkage between RNA processing and RNA translatability
A Connor1, E Wiersma, M J Shulman
1Department of Immunology, University of Toronto, Ontario, Canada.
Abstract:
The immunoglobulin mu heavy chain gene of mouse hybridoma cells is expressed in two forms, microseconds and microns, differing in their use of 3' exons. As for many other mammalian genes, mutations in the mu gene which prematurely terminate translation often have the effect of reducing the amount of these mu RNAs. To test the generality of this relationship, we selected mutant hybridoma cell lines defective in IgM production and searched both for translation termination mutations which do not reduce the amount of mu RNA as well as for mutants which show the more commonly observed reduction in mu RNA. As observed previously, the amount of microseconds RNA is normal in mutants terminating in the C mu 4 exon; by contrast the amount of microns RNA is reduced in these mutants, indicating that the effect of the mutation is influenced by some feature near the 3' end of the RNA. Mutations terminating translation in other C region exons have a graded effect on RNA content, ranging from 10% the normal level for termination in the C mu 3 exon down to 1% for termination in the C mu 2 exon. By contrast, a mutant cell line terminating in the leader exon contained 25% the normal amount of mu RNA, suggesting that translation past some point might be required to fully engage the RNA degradation process.
Insights
Investigating mutations in the mouse immunoglobulin mu heavy chain gene reveals that premature translation termination impacts RNA levels differently. Some mutations reduce RNA quantity, while others do not, depending on the mutation
Area of Science:
- Immunology
- Molecular Biology
- Genetics
Background:
- The immunoglobulin mu heavy chain gene in mouse hybridoma cells produces two forms: micro constant (µC) and micro non-polyadenylated (µN) RNAs.
- Premature translation termination mutations in mammalian genes often decrease corresponding RNA levels.
- The relationship between translation termination and RNA levels for the immunoglobulin mu heavy chain gene requires further investigation.
Purpose of the Study:
- To test the generality of the relationship between premature translation termination and RNA levels in the immunoglobulin mu heavy chain gene.
- To identify and characterize mutant hybridoma cell lines with defects in IgM production.
- To investigate mutations that either reduce or do not reduce mu RNA levels upon premature translation termination.
Main Methods:
- Selection of mutant hybridoma cell lines with impaired IgM production.
- Analysis of mu RNA levels in mutants with premature translation termination.
- Identification of mutations within specific exons (Cµ4, Cµ3, Cµ2, leader exon) of the immunoglobulin mu heavy chain gene.
Main Results:
- Mutations terminating translation in the Cµ4 exon maintained normal micro constant (µC) RNA levels but reduced micro non-polyadenylated (µN) RNA levels.
- Translation termination in other C region exons (Cµ3, Cµ2) resulted in a graded decrease in mu RNA content, from 10% to 1% of normal levels.
- A mutant terminating in the leader exon retained 25% of normal mu RNA levels, suggesting translation past a certain point may be necessary for RNA degradation.
Conclusions:
- The impact of translation termination mutations on immunoglobulin mu heavy chain RNA levels is influenced by the mutation's location, particularly near the 3' end.
- The findings suggest a complex interplay between translation and RNA stability for micro heavy chain transcripts.
- Translation progression may play a role in activating RNA degradation pathways for immunoglobulin mu heavy chain mRNA.