Evidence that PTB does not stimulate HCV IRES-driven translation.
It is now well established that Hepatitis C Virus (HCV) translation is driven by an Internal Ribosome Entry Site (IRES) resulting in cap-independent translation. Such a mechanism usually occurs with the help of IRES Associated Factors (ITAFs). Moreover, an important translational feature is likely conserved from the model of classical mRNA circularisation (5'-3' cross-talk), involving the HCV RNA highly structured 3' extremity called the 3'X region. This could bind several cellular factors and modulate the translation efficacy, at least in Rabbit Reticulocyte Lysate (RRL). In particular, polypyrimidine-binding proteins have been proposed to be potential HCV ITAFs, such as Polypyrimidine Tract Binding protein (PTB). However, contradictions still exist as to the role of PTB: its ability to bind both the HCV IRES and the 3'X region leads to the hypothesis that it could positively modulate IRES-driven translation in the presence of the X structure. Results of translational and PTB-binding studies of X mutant sequences led us to discredit PTB as protagonist of 3'X region stimulation on HCV IRES-driven translation. Moreover, competition assays of X RNA in trans on IRES-driven translation demonstrate the involvement of at least two stimulating factors and led to the conclusion that this mechanism is more complex than initially thought. Although we did not identify these factors, it is no longer doubtful that there is effectively a stimulating functional interaction between the HCV IRES and the 3'X region in RRL.
Brocard
Michèle
M
Unité Postulante Régulation de la Traduction Eucaryote et Virale, URA CNRS 1966, Institut Pasteur, 25 rue du Dr. Roux-75724, Paris cedex 15, France. mbrocard@pasteur.fr
Paulous
Sylvie
S
Komarova
Anastassia V
AV
Deveaux
Vanessa
V
Kean
Katherine M
KM
D020413:3' Untranslated Regions / Q000378:metabolism / Q000502:physiology; D020121:5' Untranslated Regions / Q000378:metabolism / Q000502:physiology; D000818:Animals; D001483:Base Sequence; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism*; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D038941:Polypyrimidine Tract-Binding Protein / Q000235:genetics / Q000502:physiology*; D011485:Protein Binding; D014176:Protein Biosynthesis / Q000235:genetics / Q000502:physiology*; D012313:RNA / Q000737:chemistry; D011817:Rabbits; D012156:Reticulocytes / Q000737:chemistry / Q000378:metabolism
D020413:3' Untranslated Regions; D020121:5' Untranslated Regions; D038941:Polypyrimidine Tract-Binding Protein; D012313:RNA