translation agency

Effect of alterations in primer binding site sequence on replication of HIV-1.
Li X; Gu Z; Wainberg MA; Parniak MA; Lady Davis Institute for Medical
December 30, 1995
Natl Conf Hum Retroviruses Relat Infect (1st). 1993 Dec 12-16;:124.

The HIV-1 replication initiation primer is tRNALys,3, a specific cellular tRNA isoacceptor selected from the more than 100 different tRNA species in the infected host cell and packaged along with viral genomic RNA during HIV assembly. Eighteen nucleotides at the 3'-terminus of the primer tRNA are annealed to a complementary primer binding site (PBS) near the 5'-end of the viral genomic RNA. The mechanisms involved in the selective packaging and annealing of tRNALys,3 to the HIV-1 PBS are unclear. In order to study the role of the PBS identity in these processes, we have mutated infectious molecular clones of HIV-1 (HXB2D and pSV21) to delete or to replace the HIV-1 PBS with sequences complementary to tRNALys,2 or tRNAPhe. Transfection of wild- type HXB2B into either Jurkat or MT-4 cells resulted in rapid emergence and spreading of cytopathic effects (CPE), but transfection of any of the mutants resulted in barely detectable CPE and yielded very low numbers of viral particles. Transfection of wild-type or mutant pSV21 constructs into COS cells produced equally high levels of virus particles. All particles showed the normal spectrum of viral proteins and genomic RNA. Infectivity of these particles was tested using MT-4 and CEM-MT-4 cells. The PBS- deleted mutant was completely non-infectious, whereas the Lys2- and Phe-PBS mutants were about 1000-fold less infectious than wild type. The implications of alterations in PBS sequence on tRNA packaging and transcription initiation will be discussed. These studies were supported by AmFAR, the American Foundation for AIDS Research.

Binding Sites Cell Line Cytopathogenic Effect, Viral *DNA Primers Human HIV-1/*PHYSIOLOGY RNA, Transfer, Lys/GENETICS Virus Replication/*GENETICS ABSTRACT