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Báo cáo y học: Contribution of the C-terminal tri-lysine regions of human immunodeficiency virus type 1 integrase for efficient reverse transcription and viral DNA nuclear import
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Tuyển tập các báo cáo nghiên cứu về y học được đăng trên tạp chí y học quốc tế cung cấp cho các bạn kiến thức về ngành y đề tài:" Contribution of the C-terminal tri-lysine regions of human immunodeficiency virus type 1 integrase for efficient reverse transcription and viral DNA nuclear import
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Báo cáo y học: " Contribution of the C-terminal tri-lysine regions of human immunodeficiency virus type 1 integrase for efficient reverse transcription and viral DNA nuclear import"Retrovirology BioMed Central Open AccessResearchContribution of the C-terminal tri-lysine regions of humanimmunodeficiency virus type 1 integrase for efficient reversetranscription and viral DNA nuclear importZhujun Ao1,2,3, Keith R Fowke2, Éric A Cohen3 and Xiaojian Yao*1,2,3Address: 1Laboratory of Molecular Human Retrovirology, Faculty of Medicine, University of Manitoba, Winnipeg, Manitoba R3E 0W3, Canada,2Department of Medical Microbiology, Faculty of Medicine, University of Manitoba, Winnipeg, Manitoba R3E 0W3, Canada and 3Laboratory ofHuman Retrovirology, Institut de Recherches Cliniques de Montréal, Département de microbiologie et immunologie, Faculté de Médecine,Université de Montréal, Montréal, Quebec H2W 1R7, CanadaEmail: Zhujun Ao - ao@cc.umanitoba.ca; Keith R Fowke - fowkekr@cc.umanitoba.ca; Éric A Cohen - Eric.Cohen@ircm.qc.ca;Xiaojian Yao* - yao2@cc.umanitoba.ca* Corresponding authorPublished: 18 October 2005 Received: 05 August 2005 Accepted: 18 October 2005Retrovirology 2005, 2:62 doi:10.1186/1742-4690-2-62This article is available from: http://www.retrovirology.com/content/2/1/62© 2005 Ao et al; licensee BioMed Central Ltd.This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0),which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: In addition to mediating the integration process, HIV-1 integrase (IN) has also been implicated in different steps during viral life cycle including reverse transcription and viral DNA nuclear import. Although the karyophilic property of HIV-1 IN has been well demonstrated using a variety of experimental approaches, the definition of domain(s) and/or motif(s) within the protein that mediate viral DNA nuclear import and its mechanism are still disputed and controversial. In this study, we performed mutagenic analyses to investigate the contribution of different regions in the C-terminal domain of HIV-1 IN to protein nuclear localization as well as their effects on virus infection. Results: Our analysis showed that replacing lysine residues in two highly conserved tri-lysine regions, which are located within previously described Region C (235WKGPAKLLWKGEGAVV) and sequence Q (211KELQKQITK) in the C-terminal domain of HIV-1 IN, impaired protein nuclear accumulation, while mutations for RK263,4 had no significant effect. Analysis of their effects on viral infection in a VSV-G pseudotyped RT/IN trans-complemented HIV-1 single cycle replication system revealed that all three C- terminal mutant viruses (KK215,9AA, KK240,4AE and RK263,4AA) exhibited more severe defect of induction of β-Gal positive cells and luciferase activity than an IN class 1 mutant D64E in HeLa-CD4- CCR5-β-Gal cells, and in dividing as well as non-dividing C8166 T cells, suggesting that some viral defects are occurring prior to viral integration. Furthermore, by analyzing viral DNA synthesis and the nucleus- associated viral DNA level, the results clearly showed that, although all three C-terminal mutants inhibited viral reverse transcription to different extents, the KK240,4AE mutant exhibited most profound effect on this step, whereas KK215,9AA significantly impaired viral DNA nuclear import. In addition, our analysis could not detect viral DNA integration in each C-terminal mutant infection, even though they displayed various low levels of nucleus-associated viral DNA, suggesting that these C-terminal mutants also impaired viral DNA integration ability. Conclusion: All of these results indicate that, in addition to being involved in HIV-1 reverse transcription and integration, the C-terminal tri-lysine regions of IN also contribute to efficient viral DNA nuclear import during the early stage of HIV-1 replication. Page 1 of ...
Nội dung trích xuất từ tài liệu:
Báo cáo y học: " Contribution of the C-terminal tri-lysine regions of human immunodeficiency virus type 1 integrase for efficient reverse transcription and viral DNA nuclear import"Retrovirology BioMed Central Open AccessResearchContribution of the C-terminal tri-lysine regions of humanimmunodeficiency virus type 1 integrase for efficient reversetranscription and viral DNA nuclear importZhujun Ao1,2,3, Keith R Fowke2, Éric A Cohen3 and Xiaojian Yao*1,2,3Address: 1Laboratory of Molecular Human Retrovirology, Faculty of Medicine, University of Manitoba, Winnipeg, Manitoba R3E 0W3, Canada,2Department of Medical Microbiology, Faculty of Medicine, University of Manitoba, Winnipeg, Manitoba R3E 0W3, Canada and 3Laboratory ofHuman Retrovirology, Institut de Recherches Cliniques de Montréal, Département de microbiologie et immunologie, Faculté de Médecine,Université de Montréal, Montréal, Quebec H2W 1R7, CanadaEmail: Zhujun Ao - ao@cc.umanitoba.ca; Keith R Fowke - fowkekr@cc.umanitoba.ca; Éric A Cohen - Eric.Cohen@ircm.qc.ca;Xiaojian Yao* - yao2@cc.umanitoba.ca* Corresponding authorPublished: 18 October 2005 Received: 05 August 2005 Accepted: 18 October 2005Retrovirology 2005, 2:62 doi:10.1186/1742-4690-2-62This article is available from: http://www.retrovirology.com/content/2/1/62© 2005 Ao et al; licensee BioMed Central Ltd.This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0),which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: In addition to mediating the integration process, HIV-1 integrase (IN) has also been implicated in different steps during viral life cycle including reverse transcription and viral DNA nuclear import. Although the karyophilic property of HIV-1 IN has been well demonstrated using a variety of experimental approaches, the definition of domain(s) and/or motif(s) within the protein that mediate viral DNA nuclear import and its mechanism are still disputed and controversial. In this study, we performed mutagenic analyses to investigate the contribution of different regions in the C-terminal domain of HIV-1 IN to protein nuclear localization as well as their effects on virus infection. Results: Our analysis showed that replacing lysine residues in two highly conserved tri-lysine regions, which are located within previously described Region C (235WKGPAKLLWKGEGAVV) and sequence Q (211KELQKQITK) in the C-terminal domain of HIV-1 IN, impaired protein nuclear accumulation, while mutations for RK263,4 had no significant effect. Analysis of their effects on viral infection in a VSV-G pseudotyped RT/IN trans-complemented HIV-1 single cycle replication system revealed that all three C- terminal mutant viruses (KK215,9AA, KK240,4AE and RK263,4AA) exhibited more severe defect of induction of β-Gal positive cells and luciferase activity than an IN class 1 mutant D64E in HeLa-CD4- CCR5-β-Gal cells, and in dividing as well as non-dividing C8166 T cells, suggesting that some viral defects are occurring prior to viral integration. Furthermore, by analyzing viral DNA synthesis and the nucleus- associated viral DNA level, the results clearly showed that, although all three C-terminal mutants inhibited viral reverse transcription to different extents, the KK240,4AE mutant exhibited most profound effect on this step, whereas KK215,9AA significantly impaired viral DNA nuclear import. In addition, our analysis could not detect viral DNA integration in each C-terminal mutant infection, even though they displayed various low levels of nucleus-associated viral DNA, suggesting that these C-terminal mutants also impaired viral DNA integration ability. Conclusion: All of these results indicate that, in addition to being involved in HIV-1 reverse transcription and integration, the C-terminal tri-lysine regions of IN also contribute to efficient viral DNA nuclear import during the early stage of HIV-1 replication. Page 1 of ...
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