Role of Mason-Pfizer Monkey Virus (MPMV) Constitutive Transport Element (CTE) in the Propagation of MPMV Vectors by Genetic Complementation Using Homologous/HeterologousenvGenes
To study Mason-Pfizer monkey virus (MPMV) replication over a single round, virus particles were generated that contain a replication-defective vector encoding a dominant selectable marker, thehygromycin B phosphotransferase (hygr)gene. Genetic complementation with a homologous MPMV envelope glycopro...
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Veröffentlicht in: | Virology (New York, N.Y.) N.Y.), 1996-10, Vol.224 (2), p.517-532 |
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Sprache: | eng |
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Zusammenfassung: | To study Mason-Pfizer monkey virus (MPMV) replication over a single round, virus particles were generated that contain a replication-defective vector encoding a dominant selectable marker, thehygromycin B phosphotransferase (hygr)gene. Genetic complementation with a homologous MPMV envelope glycoprotein (Env-gp) or pseudotyping by several heterologous Env-gps from a variety of viruses resulted in infectious MPMV particles containing the replication-defective RNA. Recently, it has been shown that human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) Rev and Rev-responsive element (RRE) functions can be substitutedin vitroby acis-acting sequence, the constitutive transport element (CTE), from simian type D retroviruses like MPMV and simian retrovirus type 1 (SRV-1). To determine whether CTE of MPMV is necessary for MPMV nucleic acid propagation, an MPMV vector that lacked the terminally located CTE was generated. Propagation of this vector was completely abrogated in the absence of CTE, showing the importance of CTE in MPMV replication. Insertion of CTE back into the MPMV genome in the sense orientation rescued replication to wild-type levels. Slot-blot analysis of nuclear versus cytoplasmic RNA fractions revealed that most of the messages were sequestered in the nucleus of cells transfected with the CTE(−) vectors and very little was transported to the cytoplasm. To test whether HIV-1 or SIV RREs could complement CTE function, the HIV-1 or SIV RREs were inserted in the CTE(−) vectors.transcomplementation of CTE(−)RRE(+) vectors with Env- and Rev- expression plasmids rescued propagation of the CTE(−) vectors. Computer analysis predicted an RNA secondary structure in MPMV CTE analogous to the HIV-1 and SIV RREs that could form three stable stem loops, the first of which contains a site similar to the Rev-binding domain in the HIV-1 RRE. The presence of a higher-order CTE structure was analyzed by mutational analysis. We conclude that CTE is important in the replication of MPMV and affects the nucleocytoplasmic transport and/or stability of viral messages similar to the Rev/RRE regulatory system of HIV-1 and SIV. |
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ISSN: | 0042-6822 1096-0341 |
DOI: | 10.1006/viro.1996.0558 |