| Summary: | <p>Abstract</p> <p>Background</p> <p>Previous studies have suggested that the process of HIV-1 tRNA primer selection and encapsidation of genomic RNA might be coupled with viral translation. In order to further investigate this relationship, proviruses were constructed in which the primer-binding site (PBS) was altered to be complementary to elongator tRNA<sup>Met </sup>(tRNA<sup>Met(e)</sup>) (HXB2-Met(e)) or initiator tRNA<sup>Met </sup>(tRNA<sup>Met(i)</sup>) (HXB2-Met(i)). These tRNA<sup>Met </sup>not only differ with respect to the 3' terminal 18-nucleotides, but also with respect to interaction with host cell proteins during protein synthesis.</p> <p>Results</p> <p>Consistent with previous studies, HXB2-Met(e) were infectious and maintained this PBS following short-term <it>in vitro </it>culture in SupT1 cells. In contrast, transfection of HBX2-Met(i) produced reduced amounts of virus (as determined by p24) and did not establish a productive infection in SupT1 cells. The low infectivity of the virus with the PBS complementary to tRNA<sup>Met(i) </sup>was not due to differences in endogenous levels of cellular tRNA<sup>Met(i) </sup>compared to tRNA<sup>Met(e)</sup>; tRNA<sup>Met(i) </sup>was also capable of being selected as the primer for reverse transcription as determined by the endogenous reverse transcription reaction. The PBS of HXB2-Met(i) contains an ATG, which could act as an upstream AUG and syphon scanning ribosomes thereby reducing initiation of translation at the authentic AUG of Gag. To investigate this possibility, a provirus with an A to G change was constructed (HXB2-Met(i)AG). Transfection of HXB2-Met(i)AG resulted in increased production of virus, similar to that for the wild type virus. In contrast to HXB2-Met(i), HXB2-Met(i)AG was able to establish a productive infection in SupT1 cells. Analysis of the PBS following replication revealed the virus favored the genome with the repaired PBS (A to G) even though tRNA<sup>Met(i) </sup>was continuously selected as the primer for reverse transcription.</p> <p>Conclusion</p> <p>The results of these studies suggest that HIV-1 has access to both tRNA<sup>Met </sup>for selection as the replication primer and supports a co-ordination between primer selection, translation and encapsidation during virus replication.</p>
|
|---|