Development and maintenance of HIV-1 latency in CD4 T cells

Paula C. Soto, UC San Diego

To date it is unclear how HIV-1 latency is established and maintained in CD4 T cells, in part due to the extremely low frequency of latently infected cells in the peripheral circulation of infected individuals. The main goal of this proposal is to identify specific species of HIV RNA that may be present in latently infected CD4 cells, and to determine their importance in maintaining HIV infection in a latent/persistent state.

To investigate cellular and viral factors involved in maintaining HIV latency, our lab has developed a unique in vitro model of HIV latency in primary CD4 T cells. This model has been optimized to yield the greatest number of viable, persistently infected CD4 cells for molecular studies of HIV latency. In order to determine which species of HIV RNA may be present in latently infected CD4 T cells, specific primer and probe sets targeting unspliced, multiply-spliced, nef-encoding, tat-encoding, and singly-spliced env-encoding species of HIV RNA have been designed. Using our in vitro model and these primers, we will determine which transcripts are present and their relative abundance by quantitative RT- PCR. We will also determine if proteins are being derived from the detected RNAs by Western blotting total protein extracted from a homogeneous population of latently infected cells. And finally, to determine if the transcripts and/or proteins identified are important for maintaining HIV in a latent state, we will disrupt their expression by using RNA interference and determine the effect of this knock-down on virus reactivation.

Upon completion of these studies we will have a better understanding of the molecular basis for viral latency. Viral factors identified have the potential to become targets of new treatment strategies to eliminate HIV-1 latent reservoirs.