REQUIREMENT FOR HIV-1 TAR SEQUENCES FOR TAT ACTIVATION IN RODENT CELLS

HIV-1 gene expression is activated via an interaction between the vira lly encoded Tat protein and a target RNA, TAR. TAR is located at the i mmediate 5' end of all viral mRNAs and comprises a partially base-pair ed stem with a tripyrimidine bulge in the upper stem and a hexanucleot ide loop. In vitro, Tat binds specifically to the bulge and upper stem region with no requirement for the loop. In contrast, when Tat activa tion is analyzed in primate cells, mutations in the loop abolish activ ation, suggesting a critical role for loop binding cellular factors. H owever, in rodent cells the reverse is true. Messages with a mutation in the TAR loop are activated whereas messages harboring a wild-type T AR sequence are not activated. By testing the effect of mutations in t he bulge and stem in the context of mutation in the loop we now show t hat this loop-independent activation by Tat in rodent cells requires t he critical bulge-stem sequences needed for Tat binding in vitro. Thes e data suggest that in rodent cells, as in vitro, Tat does not require a loop binding cofactor. In rodent cells containing human chromosome 12 (CHO12), however, Tat activation is both bulge and loop dependent. It appears that rodent cells, but not CHO12 cells, are refractory to t he normal tat/TAR activation pathway not by virtue of lacking a loop b inding cofactor, but rather by the presence of a loop binding inhibito r of either Tat binding or the activation process. (C) 1995 Academic P ress, Inc.