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Predominant usage of the proximal poly(A) site in alpha mRNAs is not intrinsic to the 3' termini
J H Coyle1, S C Borinstein, E O Woodward
1Department of Microbiology and Immunology, Virginia Commonwealth University, Richmond 23298-0678, USA.
Abstract:
The maturation of IgM-expressing B cells to IgM-secreting plasma cells is associated with both an increase in mu mRNA and the ratio of secreted to membrane forms of mu mRNA. In contrast, previous studies demonstrated that in vitro the secreted form of alpha mRNA (alpha s mRNA) predominates regardless of the stage of B cell differentiation. The present study demonstrates that alpha s mRNA predominates in both B cells derived from the germinal centers of murine Peyer's patches and in the functional IgA memory population, suggesting that in vitro events accurately represent the generation of a secretory IgA response in vivo. Although the predominant usage of the alpha s poly(A) site is due to RNA processing, it does not depend on either the alpha s poly(A) site, the 3' splice site associated with the exon encoding the membrane exon of IgA (alphaM) or the alphaM poly(A) sites. Analysis of the sequence of the intron between the alpha s terminus and alphaM (alpha s-alphaM intron) demonstrates the existence of several potential regulatory elements. Furthermore, the effects of deletions within the alpha s-alphaM intron on 3' terminus usage demonstrate that the predominant usage of the proximal terminus is not strictly dependent on the length of the intron. Together with previous work, these observations support the idea that choice of 3' terminus for all Ig heavy chain genes is regulated by a similar mechanism, but specific sequences within a heavy chain gene can impinge upon that mechanism.
Insights
Secretory IgA (sIgA) mRNA processing is consistent in vitro and in vivo, with the secreted form predominating in B cells. This suggests a conserved mechanism regulates Ig heavy chain 3' terminus selection.
Area of Science:
- Immunology
- Molecular Biology
- Genetics
Background:
- B cell differentiation involves changes in immunoglobulin (Ig) mRNA processing.
- Previous studies indicated in vitro predominant usage of secreted alpha mRNA (alpha s mRNA) across B cell stages.
- The in vivo relevance of these in vitro findings for IgA secretion was unclear.
Purpose of the Study:
- To investigate the in vivo relevance of in vitro alpha s mRNA predominance.
- To determine the regulatory mechanisms governing IgA 3' end processing in B cells.
- To compare IgA mRNA processing with that of other Ig heavy chains.
Main Methods:
- Analysis of alpha s mRNA predominance in murine Peyer's patch germinal center B cells and IgA memory B cells.
- Investigation of the role of polyadenylation (poly(A)) sites and splice sites in alpha s mRNA processing.
- Deletion analysis of the intron between alpha s and alphaM (alpha s-alphaM intron) to identify regulatory elements.
Main Results:
- Alpha s mRNA predominates in vivo in both germinal center B cells and IgA memory B cells.
- Predominant alpha s mRNA usage is RNA processing-dependent but not reliant on specific poly(A) or splice sites.
- The alpha s-alphaM intron contains potential regulatory elements, and its length is not the sole determinant of 3' terminus usage.
Conclusions:
- In vitro findings accurately reflect in vivo secretory IgA (sIgA) responses.
- A conserved mechanism likely regulates 3' terminus selection for all Ig heavy chain genes.
- Specific sequences within Ig heavy chain genes can influence this regulatory mechanism.