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J Biol Chem, Vol. 273, Issue 51, 34551-34557, December 18, 1998
From the Department of Microbiology and Program in Molecular
Biology, University of Iowa, Iowa City, Iowa 52242
Alternative splicing is used to generate more
than 30 human immunodeficiency virus type 1 (HIV-1) spliced and
unspliced mRNAs from a single primary transcript. The abundance of
HIV-1 mRNAs is determined by the efficiencies with which its
different 5' and 3' splice sites are used. Three splice sites (A4c,
A4a, and A4b) are upstream of the rev initiator AUG. RNAs
spliced at A4c, A4a, and A4b are used as mRNAs for Rev. Another 3'
splice site (A5) is immediately downstream of the rev
initiator. RNAs spliced at A5 are used as mRNAs for Env and Nef. In
this report, primer extension analysis of splicing intermediates was
used to show that there are eight branch points in this region, all of
which map to adenosine residues. In addition, cis elements recognized by the cellular splicing machinery overlap; the two most 3' branch points overlap with the AG dinucleotides at rev 3' splice
sites A4a and A4b. Competition of the overlapping cis sites for
different splicing factors may play a role in maintaining the
appropriate balance of mRNAs in HIV-1-infected cells. In support of
this possibility, mutations at rev 3' splice site A4b AG
dinucleotide dramatically increased splicing of the env/nef
3' splice site A5. This correlated with increased usage of the four
most 3' branch points, which include those within the rev
3' splice site AG dinucleotides. Consistent with these results,
analysis of a mutant in which three of the four env/nef
branch points were inactivated indicated that use of splice site A5 was
inhibited and splicing was shifted predominantly to the most 5'
rev 3' splice site A4c with preferential use of the two
most 5' branch points. Our results suggest that spliceosomes formed at
rev A4a-4b, rev A4c, and env/nef A5
3' splice sites each recognize different subsets of the eight branch
point sequences.
Overlapping Cis Sites Used for Splicing of HIV-1
env/nef and rev mRNAs
Copyright © 1998 by The American Society for Biochemistry and Molecular Biology, Inc.
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