期刊
VIROLOGY
卷 397, 期 2, 页码 346-357出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.virol.2009.11.019
关键词
Env pseudotyping; HIV-1 gene arrangement; HIV-1 Env expression; HIV-1 subtype C; HIV-1 subtype A
类别
资金
- Center for HIV/AIDS Vaccine Immunology [U01 AI067854]
- Bill & Melinda Gates Foundation [37874, 38619]
- shared core facilities of the UAB Center for AIDS Research [P30 AI27767]
- Comprehensive Cancer Center [P30 CA13148]
- Medical Scientist Training Program [T32 GM008361]
Functional Studies of HIV-1 envelope glycoproteins (Envs) commonly include the generation of pseudoviruses, which are produced by co-transfection of rev-vpu-env cassettes with an env-deficient provirus. Here, we describe six Env constructs from transmitted/founder HIV-1 that were defective in the pseudotyping assay, although two produced infectious virions when expressed from their cognate proviruses. All of these constructs exhibited an unusual gene arrangement in which the first exon of rev (rev1) and vpu were in the same reading frame without an intervening stop codon. Disruption of the rev1-vpu fusion gene by frameshift Mutation, stop codon, or abrogation of the rev initiation codon restored pseudovirion infectivity. Introduction of the fusion gene into wildtype Env cassettes severely Compromised their function. The defect was not due to altered env and rev transcription or a dominant negative effect of the expressed fusion protein, but seemed to be caused by inefficient translation at the env initiation codon. Although the rev1-vpu polymorphism affects Env expression only in vitro, it can cause problems in studies requiring Env complementation, such as analyses of co-receptor usage and neutralization properties, since 3% of subtype A, 20% of subtype C and 5% of CRF01_A/E Viruses encode the fusion gene. A solution is to eliminate the rev initiation codon when amplifying rev-vpu-env cassettes since this increases Env expression irrespective of the presence of the polymorphism. (C) 2009 Elsevier Inc. All rights reserved.
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