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Use of bi‐cistronic mRNAs, translation factors and reticulocyte lysate
Author(s) -
Merrick William C.,
Bhasker Raman C.,
Baus Diane,
Zoll Wendy L.
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a852-a
Subject(s) - internal ribosome entry site , translation (biology) , reticulocyte , eukaryotic translation , eukaryotic initiation factor , protein biosynthesis , microbiology and biotechnology , messenger rna , initiation factor , biology , eif4e , five prime untranslated region , chemistry , genetics , gene
The previous presentations by Drs. Jonathan Lorsch and Christopher Hellen have pointed out how elegantly purified translation factors can be used to assess different aspects of translation initiation. However, like most studies of complex processes with purified components, the overall rates of synthesis are reduced or quite reduced compared to in vivo rates of translation such as seen in rabbit reticulocyte lysates. We have elected to examine start site selection and IRES‐mediated initiation of a number of mRNAs as these processes are influenced by the addition of individual translation factors or by the addition of inhibitors of translation. As might be anticipated, not all the results were identical. However, two generalities do appear to be emerging from these studies. The first is that the action of eIF5 and eIF5B appear indistinguishable in start site selection and the addition of these factors tended to inhibit translation and favor initiation from the upstream start site. With respect to IRES‐mediated translation, usually addition of m 7 GTP (a cap analog) or eIF4F enhanced expression from the IRES element. A perhaps surprising result was that reduction of ternary complex levels had a much more pronounced affect on cap‐dependent translation, often with little or no reduction of expression from the IRES element.