Expression of heterologous genes from an IRES translational cassette in replication competent murine leukemia virus vectors

TY - JOUR

T1 - Expression of heterologous genes from an IRES translational cassette in replication competent murine leukemia virus vectors

AU - Jespersen, Thomas

AU - Duch, Mogens R.

AU - Carrasco, M L

AU - Warming, S

AU - Pedersen, F S

PY - 1999/11/1

Y1 - 1999/11/1

N2 - We describe replication competent retroviruses capable of expressing heterologous genes during multiple rounds of infection. An internal ribosome entry site (IRES) from encephalomyocarditis virus was inserted in the U3 region of Akv- and SL3-3-murine leukemia viruses (MLV) to direct translation of neo or the enhanced green fluorescence protein gene (EGFP). Akv-MLV's with IRES-neo and IRES-EGFP cassettes replicated with titers of about 10(6) infectious units/ml while SL3-3-MLV with IRES-neo gave about 10(3)-fold lower titers. Interestingly, RNA analysis showed a drastic reduction in the amount of spliced env mRNA for the SL3-3 derived vector relative to the Akv derived vectors, seemingly contributing to its low replication capacity. The EGFP expressing Akv-MLV was genetically stable for multiple rounds of infection; marker-cassette deletion revertants appeared after several replication rounds and these revertants only slowly became dominant in the virus population.

AB - We describe replication competent retroviruses capable of expressing heterologous genes during multiple rounds of infection. An internal ribosome entry site (IRES) from encephalomyocarditis virus was inserted in the U3 region of Akv- and SL3-3-murine leukemia viruses (MLV) to direct translation of neo or the enhanced green fluorescence protein gene (EGFP). Akv-MLV's with IRES-neo and IRES-EGFP cassettes replicated with titers of about 10(6) infectious units/ml while SL3-3-MLV with IRES-neo gave about 10(3)-fold lower titers. Interestingly, RNA analysis showed a drastic reduction in the amount of spliced env mRNA for the SL3-3 derived vector relative to the Akv derived vectors, seemingly contributing to its low replication capacity. The EGFP expressing Akv-MLV was genetically stable for multiple rounds of infection; marker-cassette deletion revertants appeared after several replication rounds and these revertants only slowly became dominant in the virus population.

KW - 3T3 Cells

KW - Animals

KW - DNA, Viral

KW - Encephalomyocarditis virus

KW - Flow Cytometry

KW - Gene Expression Regulation

KW - Genetic Vectors

KW - Green Fluorescent Proteins

KW - Kanamycin Kinase

KW - Leukemia Virus, Murine

KW - Luminescent Proteins

KW - Mice

KW - Microscopy, Fluorescence

KW - Protein Biosynthesis

KW - RNA

KW - RNA Splicing

KW - Recombinant Fusion Proteins

KW - Transformation, Genetic

KW - Virus Replication

M3 - Journal article

C2 - 10548723

SN - 0378-1119

VL - 239

SP - 227

EP - 235

JO - Gene

JF - Gene

IS - 2

ER -