Cell cycle regulation of hepatitis C and encephalomyocarditis virus internal ribosome entry site-mediated translation in human embryonic kidney 293 cells.
We have established stably transformed human embryonic kidney cell lines (HEK293) containing bicistronic constructs to study regulation of viral internal ribosome entry site (IRES)-mediated translation in vivo. These cells produce Renilla luciferase (Rluc) in a cap-dependent manner, while Firefly luciferase (Luc) synthesis is mediated by IRES elements. Using these cell lines, we demonstrate here that IRES-mediated translation directed by both hepatitis C (HCV) and encephalomyocarditis (EMCV) virus varies with the cell cycle. Experiments involving arrest of the cell lines at different phases of the cell cycle, release of synchronized cells from cell cycle arrest, as well as direct sorting of the cells based on position in the cell cycle have shown that the activity of the HCV and EMCV IRES elements is lowest during the G2/M phase in HEK293 cells. These results suggest that cellular trans-acting factors either stimulate viral IRES-mediated translation during G1 and S phases or repress translation during the G2/M phase in HEK293 cells.
Venkatesan
Arun
A
Molecular Biology Institute, University of California, Los Angeles, CA 90095, USA. dasgupta@ucla.edu
Sharma
Rakhi
R
Dasgupta
Asim
A
D020121:5' Untranslated Regions; D002453:Cell Cycle / Q000502:physiology*; D002461:Cell Line, Transformed; D004274:DNA, Recombinant; D004686:Encephalomyocarditis virus / Q000235:genetics* / Q000502:physiology; D015967:Gene Expression Regulation, Viral; D017930:Genes, Reporter; D016174:Hepacivirus / Q000235:genetics* / Q000502:physiology; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis / Q000378:metabolism; D010957:Plasmids; D014176:Protein Biosynthesis; D038621:Regulatory Sequences, Ribonucleic Acid; D014170:Transformation, Genetic
D020121:5' Untranslated Regions; D004274:DNA, Recombinant; D038621:Regulatory Sequences, Ribonucleic Acid; D008156:Luciferases