Arginine-specific regulation mediated by the Neurospora crassa arg-2 upstream open reading frame in a homologous, cell-free in vitro translation system.

The Journal of Biological Chemistry
Z Wang, Matthew S Sachs

Abstract

Translational control mediated by an upstream open reading frame (uORF) in the 5'-leader of the Neurospora crassa arg-2 mRNA was reconstituted in a homologous, cell-free in vitro translation system. A cell-free N. crassa system was developed that required the presence of cap and poly(A) on RNA for maximal translation and that was amino acid-dependent. The 24-codon arg-2 uORF, when placed in the 5'-leader region of capped and adenylated synthetic luciferase RNAs, conferred Arg-specific negative regulation in this system. Improving the uORF translation initiation context decreased luciferase production and only slightly increased the magnitude of Arg-specific regulation. Mutation of uORF Asp codon 12 to Asn, which eliminates Arg-specific regulation in vivo, eliminated regulation in vitro. Elimination of the uORF translation initiation codon also eliminated Arg-specific regulation. Arg-specific regulation in vitro appeared to be reversible. Control of RNA stability did not appear to be a primary component of Arg-specific regulation in vitro. Comparison of the effects of adding Arg to in vitro translation reactions with adding compounds related to Arg indicated that Arg-specific translational regulation was specific for L-arginine.

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Citations

Jul 24, 2002·Fungal Genetics and Biology : FG & B·Peng FangMatthew S Sachs
Oct 5, 2010·The Journal of Biological Chemistry·Christina C SpevakMatthew S Sachs
Dec 6, 2002·Nucleic Acids Research·Joseph LeeJerry Pelletier
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Apr 18, 2012·Antimicrobial Agents and Chemotherapy·Bing ZhaiXiaorong Lin
Apr 18, 2012·Molecular and Cellular Biology·Jiajie WeiMatthew S Sachs
Jan 19, 1999·Annual Review of Cell and Developmental Biology·N K Gray, M Wickens
Mar 17, 2004·Proceedings of the National Academy of Sciences of the United States of America·Peng FangMatthew S Sachs
Oct 14, 2014·Journal of Molecular Biology·Jianli Lu, Carol Deutsch
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Aug 11, 2016·Proceedings of the National Academy of Sciences of the United States of America·Stephen Z CasterDeborah Bell-Pedersen
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