Impact of human immunodeficiency virus type 1 RNA dimerization on viral infectivity and of stem-loop B on RNA dimerization and reverse transcription and dissociation of dimerization from packaging.
The kissing-loop domain (KLD) encompasses a stem-loop, named kissing-loop or dimerization initiation site (DIS) hairpin (nucleotides [nt] 248 to 270 in the human immunodeficiency virus type 1 strains HIV-1(Lai) and HIV-1(Hxb2)), seated on top of a 12-nt stem-internal loop called stem-loop B (nt 243 to 247 and 271 to 277). Destroying stem-loop B reduced genome dimerization by approximately 50% and proviral DNA synthesis by approximately 85% and left unchanged the dissociation temperature of dimeric genomic RNA. The most affected step of reverse transcription was plus-strand DNA transfer, which was reduced by approximately 80%. Deleting nt 241 to 256 or 200 to 256 did not reduce genome dimerization significantly more than the destruction of stem-loop B or the DIS hairpin. We conclude that the KLD is nonmodular: mutations in stem-loop B and in the DIS hairpin have similar effects on genome dimerization, reverse transcription, and encapsidation and are also "nonadditive"; i.e., a larger deletion spanning both of these structures has the same effects on genome dimerization and encapsidation as if stem-loop B strongly impacted DIS hairpin function and vice versa. A C258G transversion in the palindrome of the kissing-loop reduced genome dimerization by approximately 50% and viral infectivity by approximately 1.4 log. Two mutations, CGCG261-->UUAA261 (creating a weaker palindrome) and a Delta241-256 suppressor mutation, were each able to reduce genome dimerization but leave genome packaging unaffected.
Shen
N
N
McGill AIDS Centre, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis-Jewish General Hospital, McGill University, Montreal, Quebec, Canada H3T 1E2.
Jetté
L
L
Liang
C
C
Wainberg
M A
MA
Laughrea
M
M
D000818:Animals; D020029:Base Pairing / Q000235:genetics; D001665:Binding Sites; D002213:Capsid / Q000235:genetics; D036022:Capsid Proteins; D002460:Cell Line; D004279:DNA, Viral / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D019281:Dimerization; D015683:Gene Products, gag / Q000235:genetics; D016153:Genes, Suppressor / Q000235:genetics; D016679:Genome, Viral; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology*; D006801:Humans; D008954:Models, Biological; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D010446:Peptide Fragments / Q000235:genetics; D011533:Proviruses / Q000235:genetics; D020871:RNA Stability; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D014158:Transcription, Genetic; D014764:Viral Proteins; D019065:Virus Assembly; D054301:gag Gene Products, Human Immunodeficiency Virus
D036022:Capsid Proteins; D004279:DNA, Viral; D015683:Gene Products, gag; C071884:NCP7 protein, Human immunodeficiency virus 1; D010446:Peptide Fragments; D012367:RNA, Viral; D014764:Viral Proteins; D054301:gag Gene Products, Human Immunodeficiency Virus; C109809:p2 gag peptide, Human immunodeficiency virus 1