Alternative splice acceptor utilization during human immunodeficiency virus type 1 infection of cultured cells.
The utilization of alternative splice acceptors for excision of the 5' major intron of human immunodeficiency virus type 1 RNA was observed after infection in vitro. Specific splice events were monitored by a cDNA-polymerase chain reaction. These splice events shared a common splice donor but utilized several alternative splice acceptors. In addition to identifying the previously documented splice acceptors for tat and nef (S. K. Arya, C. Guo, S. F. Josephs, and F. Wong-Staal, Science 229:69-73, 1985), nucleotide sequence analysis of cDNA-polymerase chain reaction fragments also revealed the following: (i) two splice acceptors 15 and 9 nucleotides upstream from the rev start codon, which are utilized to create transcripts suitable for specific rev expression; and (ii) use of the splice acceptor previously attributed to nef to generate a singly spliced, env-encoding transcript. Hybridization signals representing the nef/env, tat, and rev splice events increased in intensity between 6 and 12 h after infection of CEM cells with the LAV-1BRU strain of human immunodeficiency virus type 1. In contrast, the signal for utilization of the nef/env splice acceptor for the singly spliced env transcript appeared first at 12 h and increased to maximum intensity by 24 h. The nef/env splice acceptor was dominant at all time points examined. We propose that this dominance ensures efficient downstream splicing proximal to the env initiation codon in singly spliced transcripts. However, early after infection, the dominance of the nef/env splice acceptor appears to divert primary transcripts away from tat- and rev-specific processing paths. The relative proportions of hybridization signals representing these alternative splice events remained constant throughout the viral replicative cycle. This result suggests that trans-acting factors that might influence splice choices are not induced during infection, but rather that cis-acting, sequence-specific splice preferences determine the relative efficiency of alternative acceptor utilization.
Guatelli
J C
JC
Department of Medicine, University of California, San Diego.
Gingeras
T R
TR
Richman
D D
DD
D001483:Base Sequence; D002460:Cell Line; D002472:Cell Transformation, Viral; D004279:DNA, Viral / Q000235:genetics; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D007438:Introns; D007700:Kinetics; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D012326:RNA Splicing; D012367:RNA, Viral / Q000235:genetics*
D004279:DNA, Viral; D015345:Oligonucleotide Probes; D012367:RNA, Viral