TY - JOUR
T1 - Contribution of the C-terminal region within the catalytic core domain of HIV-1 integrase to yeast lethality, chromatin binding and viral replication
AU - Xu, Zaikun
AU - Zheng, Yingfeng
AU - Ao, Zhujun
AU - Clement, Martin
AU - Mouland, Andrew J.
AU - Kalpana, Ganjam V.
AU - Belhumeur, Pierre
AU - Cohen, Éric A.
AU - Yao, Xiao Jian
N1 - Funding Information:
We thank Dr. D. Grandgenett and Dr. A. Engelman for providing the anti-IN antiserum and pFT-1-LEDGF through the AIDS Research Reference Reagent Program, Division of AIDS, NIAID, NIH. We thank Dr. Keith Fowke and Mr. John Rutherford for their technical support. YF. Zheng is a recipient of an MHRC/MICH scholarship. A.J. Mouland is a recipient of a Canadian Institutes of Health Research (CIHR) New Investigator Award and E.A. Cohen is a recipient of the Canada Research Chair in Human Ret-rovirology. X-J. Yao is a recipient of the Basic Science Career Development Research Award from The Manitoba Medical Service Foundation. This work was supported by CIHR grants (HOP-63013 and HOP-81180) and the grant from The Manitoba Medical Service Foundation to X-J. Yao.
PY - 2008
Y1 - 2008
N2 - Background: HIV-1 integrase (IN) is a key viral enzymatic molecule required for the integration of the viral cDNA into the genome. Additionally, HIV-1 IN has been shown to play important roles in several other steps during the viral life cycle, including reverse transcription, nuclear import and chromatin targeting. Interestingly, previous studies have demonstrated that the expression of HIV-1 IN induces the lethal phenotype in some strains of Saccharomyces cerevisiae. In this study, we performed mutagenic analyses of the C-terminal region of the catalytic core domain of HIV-1 IN in order to delineate the critical amino acid(s) and/or motif(s) required for the induction of the lethal phenotype in the yeast strain HP16, and to further elucidate the molecular mechanism which causes this phenotype. Results: Our study identified three HIV-1 IN mutants, V165A, A179P and KR186,7AA, located in the C-terminal region of the catalytic core domain of IN that do not induce the lethal phenotype in yeast. Chromatin binding assays in yeast and mammalian cells demonstrated that these IN mutants were impaired for the ability to bind chromatin. Additionally, we determined that while these IN mutants failed to interact with LEDGF/ p75, they retained the ability to bind Integrase interactor 1. Furthermore, we observed that VSV-G-pseudotyped HIV-1 containing these IN mutants was unable to replicate in the C8166 T cell line and this defect was partially rescued by complementation with the catalytically inactive D64E IN mutant. Conclusion: Overall, this study demonstrates that three mutations located in the C-terminal region of the catalytic core domain of HIV-1 IN inhibit the IN-induced lethal phenotype in yeast by inhibiting the binding of IN to the host chromatin. These results demonstrate that the C-terminal region of the catalytic core domain of HIV-1 IN is important for binding to host chromatin and is crucial for both viral replication and the promotion of the IN-induced lethal phenotype in yeast.
AB - Background: HIV-1 integrase (IN) is a key viral enzymatic molecule required for the integration of the viral cDNA into the genome. Additionally, HIV-1 IN has been shown to play important roles in several other steps during the viral life cycle, including reverse transcription, nuclear import and chromatin targeting. Interestingly, previous studies have demonstrated that the expression of HIV-1 IN induces the lethal phenotype in some strains of Saccharomyces cerevisiae. In this study, we performed mutagenic analyses of the C-terminal region of the catalytic core domain of HIV-1 IN in order to delineate the critical amino acid(s) and/or motif(s) required for the induction of the lethal phenotype in the yeast strain HP16, and to further elucidate the molecular mechanism which causes this phenotype. Results: Our study identified three HIV-1 IN mutants, V165A, A179P and KR186,7AA, located in the C-terminal region of the catalytic core domain of IN that do not induce the lethal phenotype in yeast. Chromatin binding assays in yeast and mammalian cells demonstrated that these IN mutants were impaired for the ability to bind chromatin. Additionally, we determined that while these IN mutants failed to interact with LEDGF/ p75, they retained the ability to bind Integrase interactor 1. Furthermore, we observed that VSV-G-pseudotyped HIV-1 containing these IN mutants was unable to replicate in the C8166 T cell line and this defect was partially rescued by complementation with the catalytically inactive D64E IN mutant. Conclusion: Overall, this study demonstrates that three mutations located in the C-terminal region of the catalytic core domain of HIV-1 IN inhibit the IN-induced lethal phenotype in yeast by inhibiting the binding of IN to the host chromatin. These results demonstrate that the C-terminal region of the catalytic core domain of HIV-1 IN is important for binding to host chromatin and is crucial for both viral replication and the promotion of the IN-induced lethal phenotype in yeast.
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U2 - 10.1186/1742-4690-5-102
DO - 10.1186/1742-4690-5-102
M3 - Article
C2 - 19014595
AN - SCOPUS:58149529842
SN - 1742-4690
VL - 5
JO - Retrovirology
JF - Retrovirology
M1 - 102
ER -
