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Related Experiment Videos

Complex alternative RNA processing generates an unexpected diversity of poly(A) polymerase isoforms

W Zhao1, J L Manley

  • 1Department of Biological Sciences, Columbia University, New York, NY 10027, USA.

Molecular and Cellular Biology
|May 1, 1996
PubMed
Summary

Multiple poly(A) polymerase (PAPs) forms are generated through complex gene expression. Short PAP forms, though transcribed, appear inactive, suggesting regulation at the translation level.

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Area of Science:

  • Molecular Biology
  • Gene Expression Regulation
  • Enzymology

Background:

  • Multiple poly(A) polymerase (PAPs) forms (I, II, III) have been identified across species.
  • Long PAP forms (I, II) possess distinct functional domains, while PAP III is truncated.
  • Previous studies characterized PAP cDNA from various organisms, but mouse gene structure and expression remained unclear.

Purpose of the Study:

  • To investigate the structure and expression of mouse poly(A) polymerase (PAPs) gene.
  • To elucidate the mechanisms generating different PAP isoforms.
  • To determine the functional activity of various PAP forms.

Main Methods:

  • Isolation of mouse PAP gene and pseudogene from a genomic library.
  • Analysis of alternative splicing and polyadenylation events.

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  • RNase protection and RT-PCR for expression analysis.
  • Immunoblot analysis and in vitro activity assays of recombinant PAPs.

    • Main Results:

    • Mouse PAP gene structure suggests alternative splicing and polyadenylation generate different PAP forms (I-VI).
    • PAP II, V, and VI are ubiquitously expressed, while PAP I, IV, and III show tissue-specific expression.
    • Immunoblotting detected only long PAP forms, and in vitro assays confirmed short PAP forms are inactive.

    Conclusions:

    • Complex gene expression mechanisms generate diverse PAP isoforms in mice.
    • Tissue-specific expression and translational regulation may control PAP function.
    • Inactive short PAP forms suggest a novel layer of post-transcriptional regulation in polyadenylation.