Viruses and Designed Host Cells

Improved assays were feasible with increased understanding of molecular events in the infectious cycle: upon entry into the cell, the viral genomic RNA molecule is reverse-transcribed into DNA, and this copy is integrated into the host chromosome as a provirus. Integration ofthe proviral DNA is the main mechanism of retroviruses to establish a persistent infection, as cell division maintains a pool of infected cells even in the absence of actively replicating virus. Induction of the HIV provirus towards production and release ofvirus particles is initiated by the viral Tat (transactivator oftranscription) protein. Because Tat is unique to HIV and pivotal in productive infectious cycles, it has been appreciated very early as a means towards providing robust assays in the search for antivirals [63, 64]. The LTR, an important regulatory and Tat-responsive stretch of proviral DNA, was fused to the gene for a reporter (lacZ or CAT), and this cassette was stably inserted into cells susceptible to HIV infection. Upon productive infection with HIV, the viral Tat protein activates reporter gene expression, and this event can be quantified in the presence or absence of proposed inhibitor molecules, for example the venerable AZT.

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