You are not logged in.
Openly accessible

Human immunodeficiency virus type 1 protease regulation of tat activity is essential for efficient reverse transcription and replication

Apolloni, Ann, Hooker, C. William, Mak, Johnson and Harrich, David 2003, Human immunodeficiency virus type 1 protease regulation of tat activity is essential for efficient reverse transcription and replication, Journal of virology, vol. 77, no. 18, pp. 9912-9921, doi: 10.1128/JVI.77.18.9912-9921.2003.

Attached Files
Name Description MIMEType Size Downloads
mak-humanimmunodeficiency-2003.pdf Published version application/pdf 897.86KB 44

Title Human immunodeficiency virus type 1 protease regulation of tat activity is essential for efficient reverse transcription and replication
Author(s) Apolloni, Ann
Hooker, C. William
Mak, JohnsonORCID iD for Mak, Johnson orcid.org/0000-0002-5229-5707
Harrich, David
Journal name Journal of virology
Volume number 77
Issue number 18
Start page 9912
End page 9921
Total pages 10
Publisher American Society for Microbiology
Place of publication Washington, D. C.
Publication date 2003-09
ISSN 0022-538X
1098-5514
Keyword(s) animals
cell Line
gene products
tat
HIV protease
HIV-1
humans
rabbits
reticulocytes
genetic transcription
virus replication
Summary The human immunodeficiency virus type 1 (HIV-1) Tat protein enhances reverse transcription, but it is not known whether Tat acts directly on the reverse transcription complex or through indirect mechanisms. Since processing of Tat by HIV protease (PR) might mask its presence and, at least in part, explain this lack of data, we asked whether Tat can be cleaved by PR. We used a rabbit reticulocyte lysate (RRL) system to make Tat and PR. HIV-1 PR is expressed as a Gag-Pol fusion protein, and a PR-inactivated Gag-Pol is also expressed as a control. We showed that Tat is specifically cleaved in the presence of PR, producing a protein of approximately 5 kDa. This result suggested that the cleavage site was located in or near the Tat basic domain (amino acids 49 to 57), which we have previously shown to be important in reverse transcription. We created a panel of alanine-scanning mutations from amino acids 45 to 54 in Tat and evaluated functional parameters, including transactivation, reverse transcription, and cleavage by HIV-1 PR. We showed that amino acids 49 to 52 (RKKR) are absolutely required for Tat function in reverse transcription, that mutation of this domain blocks cleavage by HIV-1 PR, and that other pairwise mutations in this region modulate reverse transcription and proteolysis in strikingly similar degrees. Mutation of Tat Y47G48 to AA also down-regulated Tat-stimulated reverse transcription but had little effect on transactivation or proteolysis by HIV PR, suggesting that Y47 is critical for reverse transcription. We altered the tat gene of the laboratory strain NL4-3 to Y47D and Y47N so that overlapping reading frames were not affected and showed that Y47D greatly diminished virus replication and conveyed a reverse transcription defect. We hypothesize that a novel, cleaved form of Tat is present in the virion and that it requires Y47 for its role in support of efficient reverse transcription.
Language eng
DOI 10.1128/JVI.77.18.9912-9921.2003
Field of Research 119999 Medical and Health Sciences not elsewhere classified
Socio Economic Objective 970111 Expanding Knowledge in the Medical and Health Sciences
HERDC Research category C1.1 Refereed article in a scholarly journal
Copyright notice ©2003, American Society for Microbiology
Free to Read? Yes
Persistent URL http://hdl.handle.net/10536/DRO/DU:30047537

Document type: Journal Article
Collections: School of Medicine
Open Access Collection
Connect to link resolver
 
Unless expressly stated otherwise, the copyright for items in DRO is owned by the author, with all rights reserved.

Every reasonable effort has been made to ensure that permission has been obtained for items included in DRO. If you believe that your rights have been infringed by this repository, please contact drosupport@deakin.edu.au.

Versions
Version Filter Type
Citation counts: TR Web of Science Citation Count  Cited 23 times in TR Web of Science
Scopus Citation Count Cited 27 times in Scopus
Google Scholar Search Google Scholar
Access Statistics: 203 Abstract Views, 45 File Downloads  -  Detailed Statistics
Created: Thu, 30 Aug 2012, 09:41:00 EST

Every reasonable effort has been made to ensure that permission has been obtained for items included in DRO. If you believe that your rights have been infringed by this repository, please contact drosupport@deakin.edu.au.