Deficient dimerization of human immunodeficiency virus type 1 RNA caused by mutations of the u5 RNA sequences.
The human immunodeficiency virus type 1 (HIV-1) virion contains two copies of genomic RNA that are noncovalently attached along a region at their 5' ends, in which two contact sites have been observed by electron microscopy. One of these sites is believed to be the stem-loop 1 (SL1) sequence which serves as the dimerization initiation site (DIS), and the other site, closer to the 5' end of the viral RNA, may involve the R or U5 RNA sequences. In this study, we present biochemical evidence showing that alteration of the U5 RNA sequence in the context of full-length viral RNA leads to diminished dimerization of virion RNA. In particular, two stretches of GU-rich sequences, which are located at nucleotides (nt) 99 to 108 and nt 112 to 123 within U5, were either deleted or substituted with exogenous sequences. The mutated viruses thus generated all exhibited deficient RNA dimerization. This dimerization deficit was not corrected by second-site mutations that preserved local RNA structures, such as the poly(A) hairpin, and was overcome to only a limited extent by compensatory mutations within Gag; these mutations were identified after long-term culture of the relevant mutant viruses in permissive cell lines and were able to restore viral infectiousness and RNA packaging to wild-type levels. Therefore, these GU sequences do not regulate RNA dimerization by the formation of local secondary structures nor by the maintenance of efficient viral RNA packaging; instead, they may mediate direct RNA-RNA interactions in the dimer structure. In contrast, mutation of palindrome 5'-AAGCUU-3', which resides within R and crowns the poly(A) hairpin, did not affect either RNA dimerization or RNA packaging.
Russell
Rodney S
RS
McGill AIDS Centre, Lady Davis Institute-Jewish General Hospital, Montreal, Quebec, H3T 1E2, Canada.
Hu
Jing
J
Laughrea
Michael
M
Wainberg
Mark A
MA
Liang
Chen
C
D020121:5' Untranslated Regions / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D019281:Dimerization; D015683:Gene Products, gag / Q000235:genetics; D015497:HIV-1 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D012367:RNA, Viral / Q000737:chemistry; D014779:Virus Replication
D020121:5' Untranslated Regions; D015683:Gene Products, gag; D012367:RNA, Viral