In vivo binding of wild-type and mutant human immunodeficiency virus type 1 Rev proteins: implications for function.
The Rev transactivator protein of human immunodeficiency virus type 1 (HIV-1) is required for protein expression from the HIV-1 RNAs which contain a binding site for the Rev protein, termed the Rev-responsive element (RRE). This transactivator acts both at the level of splicing/transport of nuclear RNAs and at the level of translation of cytoplasmic RNAs. We used a monoclonal antibody specific for the HIV-1 Rev protein to immunoprecipitate cellular extracts from HIV-1-infected and -transfected cells. High levels of specific binding of wild-type Rev to the RRE-containing RNAs were found in cytoplasmic, but not nuclear, extracts from these cells. A Rev mutant which lacked both nuclear and cytoplasmic Rev function but retained RNA binding in vivo was generated. This binding was detectable with both nuclear and cytoplasmic extracts. These results verify the existence of direct binding of Rev to HIV-1 RNAs in vivo and conclusively prove that binding of Rev is not sufficient for nuclear or cytoplasmic Rev function. The results also support a direct role for Rev in the nuclear export and translation of HIV-1 RNAs.
Arrigo
S J
SJ
Department of Microbiology and Immunology, Medical University of South Carolina, Charleston 29425-2230.
Heaphy
S
S
Haines
J K
JK
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D002478:Cells, Cultured; D003593:Cytoplasm / Q000737:chemistry; D015694:Gene Products, rev / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D015658:HIV Infections / Q000378:metabolism; D015497:HIV-1 / Q000378:metabolism*; D006801:Humans; D008214:Lymphocytes; D008969:Molecular Sequence Data; D016296:Mutagenesis; D012367:RNA, Viral / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D054321:rev Gene Products, Human Immunodeficiency Virus
D000911:Antibodies, Monoclonal; D015694:Gene Products, rev; D012367:RNA, Viral; D016601:RNA-Binding Proteins; D054321:rev Gene Products, Human Immunodeficiency Virus