Toeprint analysis of the positioning of translation apparatus components at initiation and termination codons of fungal mRNAs
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Authors
Sachs, Matthew S.Wang, Zhong
Gaba, Anthony
Fang, Peng
Belk, Jonathan Philip
Ganesan, Robin
Amrani, Nadia
Jacobson, Allan
UMass Chan Affiliations
Department of Molecular Genetics and MicrobiologyDepartment of Biochemistry and Molecular Biology
Graduate School of Biomedical Sciences
Document Type
Journal ArticlePublication Date
2002-06-11Keywords
Base Sequence; Chromatography; Codon, Terminator; Genes, Fungal; Molecular Sequence Data; Neurospora crassa; Nitrogen; Open Reading Frames; *Protein Biosynthesis; RNA, Messenger; Saccharomyces cerevisiae; TemperatureLife Sciences
Medicine and Health Sciences
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Show full item recordAbstract
The ability to map the position of ribosomes and their associated factors on mRNAs is critical for an understanding of translation mechanisms. Earlier approaches to monitoring these important cellular events characterized nucleotide sequences rendered nuclease-resistant by ribosome binding. While these approaches furthered our understanding of translation initiation and ribosome pausing, the pertinent techniques were technically challenging and not widely applied. Here we describe an alternative assay for determining the mRNA sites at which ribosomes or other factors are bound. This approach uses primer extension inhibition, or "toeprinting," to map the 3' boundaries of mRNA-associated complexes. This methodology, previously used to characterize initiation mechanisms in prokaryotic and eukaryotic systems, is used here to gain an understanding of two interesting translational regulatory phenomena in the fungi Neurospora crassa and Saccharomyces cerevisiae: (a) regulation of translation in response to arginine concentration by an evolutionarily conserved upstream open reading frame, and (b) atypical termination events that occur as a consequence of the presence of premature stop codons.Source
Methods. 2002 Feb;26(2):105-14. Link to article on publisher's siteDOI
10.1016/S1046-2023(02)00013-0Permanent Link to this Item
http://hdl.handle.net/20.500.14038/32493PubMed ID
12054887Related Resources
Link to Article in PubMedae974a485f413a2113503eed53cd6c53
10.1016/S1046-2023(02)00013-0