THE ASSOCIATION OF HIV-1 TAT WITH NUCLEI IS REGULATED BY CA2+ IONS AND CYTOSOLIC FACTORS

Human immunodeficiency virus-1 (HIV-1) Tat, a nuclear transcription fa ctor, has been shown to function extracellularly, implying that some T at molecules escape nuclear import and are secreted. This raises the q uestion of what regulates, in HIV-1-infected cells, the nuclear target ing of the polypeptide. Here we show that cytosolic components activat ed by Ca2+ ions are required to reveal the karyophilic properties of T at: in vitro translated Tat molecules do not associate with isolated n uclei unless preincubated with Ca2+. Moreover, Ca2+ ions induce karyop hilicity of chemically synthesized Tat molecules only upon addition of cytosolic extracts. The Ca2+-induced karyophilicity is prevented by i nhibitors of either tyrosine kinases (herbimycin A and genistein) or t yrosine phosphatases (vanadate), suggesting the involvement of Ca2+-de pendent phosphorylation/dephosphorylation events. In line with these o bservations, the transcriptional activity of Tat is inhibited by treat ment with either vanadate or genistein. The same occurs with Tat mutan ts lacking either one or both the two tyrosine residues (positions 26 and 47). Hence, Ca2+-dependent tyrosine kinase(s) and phosphatase(s) a ct on accessory cellular protein(s), which in turn are responsible of Tat karyophilicity.