Identification of a cellular protein that specifically interacts with the essential cysteine region of the HIV-1 Tat transactivator.
The Tat protein of the human immunodeficiency virus (HIV) is a powerful activator of HIV gene expression. Genetic and biochemical evidence suggests that one or more cellular cofactors may be important for Tat activity. We have used two-hybrid interactive cloning in yeast to identify a partial cDNA clone (clone 10) from a human B-lymphoblastoid library that specifically interacts with the N-terminal 31 amino acids of HIV-1 Tat which contains the essential cysteine-rich portion of the Tat activation domain. The encoded protein also binds to purified Tat in vitro. Mutation of single essential cysteine residues in Tat abolishes interaction between Tat and clone 10, suggesting that interaction with the encoded protein is important for Tat activity. We have identified the full-length cDNA for the Tat binding protein and shown that overexpression of the encoded protein, Tip60 (Tat interactive protein, 60 kDa), results in a fourfold augmentation of Tat transactivation of the HIV-1 promoter in transient expression assays without increasing the basal activity of the HIV promoter or activating the heterologous RSV promoter. These data together with the genetic and in vitro binding data support the notion that Tip60 might be a cofactor of Tat involved in the regulation of HIV gene expression.
Kamine
J
J
Institute for Molecular Virology, St. Louis University School of Medicine, Missouri 63110, USA.
Elangovan
B
B
Subramanian
T
T
Coleman
D
D
Chinnadurai
G
G
D000123:Acetyltransferases; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003545:Cysteine / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D051548:Histone Acetyltransferases; D006801:Humans; D000074443:Lysine Acetyltransferase 5; D008969:Molecular Sequence Data; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011506:Proteins / Q000235:genetics*; D012441:Saccharomyces cerevisiae; D054322:tat Gene Products, Human Immunodeficiency Virus
D018076:DNA, Complementary; D015696:Gene Products, tat; D011506:Proteins; D054322:tat Gene Products, Human Immunodeficiency Virus; D000123:Acetyltransferases; D051548:Histone Acetyltransferases; C098489:KAT5 protein, human; D000074443:Lysine Acetyltransferase 5; D003545:Cysteine