This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Prof. Charles P. Scholes from Albany University is actively studying the mechanisms underlying HIV-1 replication. A critical step in HIV-1 replication is reverse transcription of HIV-1 genomic RNA to double-stranded DNA by reverse transcriptase (RT). Two viral proteins, the RT enzyme and a small nucleocapsid protein NCp7, which is well-known nucleic acid chaperone direct this process. At the beginning of the linear DNA synthesis, the newly made minus-strand strong-stop DNA ((-)ssDNA) is transferred to the 3'end of the genomic RNA by means of an hybridization reaction between transactivation response element (TAR) RNA and cTAR DNA sequences. Since both TAR sequences exhibit stable hairpin structures, NCp7 needs to destabilize the TAR structures in order to chaperone their hybridization. NCp7 is characterized by two zinc fingers that plays a key role in HIV-1 life cycle and presents a promising target for an antiviral therapy. It has been shown that NCp7 chaperones the first strand transfer by promoting the annealing of the complementary transactivation response element (TAR) RNA and cTAR DNA stem-loop sequences. This critically relies on NCp7 ability to transiently melt their terminal base pairs. The destabilization activity of NCp7 is mediated by a hydrophobic plateau at the surface of the folded zinc fingers, which restricted the oligonucleotide flexibility. The destabilizing of the TAR and cTAR sequences can be mediated by a single finger motif, while the annealing activity requires the two fingers. NCp7 induces a limited destabilization of the primer binding site (PBS) stem-loops involved in the second strand transfer. However, NCp7 can chaperone their homodimerization, by stabilizing the annealing of their partly self-complementary loops. The propensity of NCp7 to promote the dimerization of partly complementary sequences may favor secondary contacts between viral sequences and thus, recombination and viral diversity. Moreover, NCp7 promotes the second strand transfer, by chaperoning a kissing-loop intermediate, through an extension of the loops and a weakening of the upper base pair of the stem.
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