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Trypanosoma brucei poly(A) binding protein I cDNA cloning, expression, and binding to 5 untranslated region sequence
T L Hotchkiss1, G E Nerantzakis, S C Dills
1Department of Microbiology and Center for Microbial Pathogenesis, SUNY Buffalo School of Medicine, NY 14214, USA.
Molecular and Biochemical Parasitology
|February 24, 1999
Summary
Poly(A) binding protein I (PABPI) in Trypanosoma brucei binds its own 5' untranslated region, suggesting autoregulation. This binding involves the A-rich sequence, impacting gene expression.
Area of Science:
- Molecular Biology
- Gene Regulation
- Parasitology
Background:
- Poly(A) binding protein I (PABPI) is crucial for mRNA processing and translation in eukaryotes.
- Investigating PABPI in Trypanosoma brucei provides insights into posttranscriptional gene regulation in this parasite.
Purpose of the Study:
- To clone and characterize the PABPI gene from Trypanosoma brucei.
- To investigate the potential autoregulatory role of T. brucei PABPI.
Main Methods:
- Cloning of PABPI cDNA from Trypanosoma brucei.
- Northern blot analysis to determine mRNA characteristics.
- Expression of PABPI as a glutathione-S-transferase fusion protein.
- UV cross-linking and competition assays to study RNA binding.
Main Results:
- The cloned cDNA encodes a protein homologous to eukaryotic PABPI, with conserved RNA binding domains.
- T. brucei PABPI exhibits an unusually long 3' untranslated region (~3 kb).
- T. brucei PABPI binds to its own 5' untranslated region, particularly the A-rich sequence, suggesting autoregulation.
Conclusions:
- T. brucei PABPI shares conserved features with orthologs from other species.
- The A-rich region of the 5' UTR is critical for PABPI autoregulation.
- PABPI may autoregulate its own expression in Trypanosoma brucei.