AN RGD CONTAINING PEPTIDE FROM HIV-1 TAT-(65-80) MODULATES PROTOONCOGENE EXPRESSION IN HUMAN BRONCHOALVEOLAR CARCINOMA CELL-LINE, A549

Tat (transactivator of transcription) is essential for HIV-1 replicati on in vivo and in vitro. Tat-(65-80), an RGD containing domain, has be en shown to regulate proliferative function of a variety of cell lines , including a human adenocarcinoma cell line, A549. The exact cellular and molecular mechanisms by which these effects are mediated, remain unknown. To evaluate the hypothesis that Tat-(65-80) modulates the exp ression of immediate early genes (IEG) c-jun, c-myc, c-fos and the tum or suppressor gene p53, serum starved A549 cells were incubated with T at-(65-80) or heat-inactivated Tat-(65-80) at 10 ng/ml. Total cellular RNA was Isolated from the cells at various time points (0-24 hours). In each case, 5 mu g of RNA was reverse transcribed in 20 mu 1 of reac tion volume. Equal amounts of cDNA were subjected to polymerase chain reaction (PCR) and analyzed by electrophoresis. The photographic negat ives of the ethidium bromide stained gels were quantitated by densitom etric scanning and normalized to corresponding beta-actin PCR products . Treatment with Tat-(65-80) showed a twofold induction of c-jun at 0. 5 h. Peak expression occurred at 60 minutes and remained above baselin e at 24 hours (h). c-myc was increased at 0.5 h, reached a twofold inc rease at 2 h and remained above baseline at 24 h. c-Sos increased seve n fold at 0.5 h and declined subsequently to baseline at 8 h, p-53 gen e was reduced fivefold at 0.5 h and remained downregulated thereafter. These results show that Tat-(65-80) can modulate growth related genes in human lung epithelial cells.