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Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles
It remains unclear what determines the subcellular localization of hepatitis B virus (HBV) core protein (HBc) and particles. To address this fundamental issue, we have identified four distinct HBc localization signals in the arginine rich domain (ARD) of HBc, using immunofluorescence confocal micros...
Autores principales: | , , , , , , , |
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Formato: | Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2010
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2965763/ https://www.ncbi.nlm.nih.gov/pubmed/21060813 http://dx.doi.org/10.1371/journal.ppat.1001162 |
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author | Li, Hung-Cheng Huang, Er-Yi Su, Pei-Yi Wu, Szu-Yao Yang, Ching-Chun Lin, Young-Sun Chang, Wen-Chang Shih, Chiaho |
author_facet | Li, Hung-Cheng Huang, Er-Yi Su, Pei-Yi Wu, Szu-Yao Yang, Ching-Chun Lin, Young-Sun Chang, Wen-Chang Shih, Chiaho |
author_sort | Li, Hung-Cheng |
collection | PubMed |
description | It remains unclear what determines the subcellular localization of hepatitis B virus (HBV) core protein (HBc) and particles. To address this fundamental issue, we have identified four distinct HBc localization signals in the arginine rich domain (ARD) of HBc, using immunofluorescence confocal microscopy and fractionation/Western blot analysis. ARD consists of four tight clustering arginine-rich subdomains. ARD-I and ARD-III are associated with two co-dependent nuclear localization signals (NLS), while ARD-II and ARD-IV behave like two independent nuclear export signals (NES). This conclusion is based on five independent lines of experimental evidence: i) Using an HBV replication system in hepatoma cells, we demonstrated in a double-blind manner that only the HBc of mutant ARD-II+IV, among a total of 15 ARD mutants, can predominantly localize to the nucleus. ii) These results were confirmed using a chimera reporter system by placing mutant or wild type HBc trafficking signals in the heterologous context of SV40 large T antigen (LT). iii) By a heterokaryon or homokaryon analysis, the fusion protein of SV40 LT-HBc ARD appeared to transport from nuclei of transfected donor cells to nuclei of recipient cells, suggesting the existence of an NES in HBc ARD. This putative NES is leptomycin B resistant. iv) We demonstrated by co-immunoprecipitation that HBc ARD can physically interact with a cellular factor TAP/NXF1 (Tip-associated protein/nuclear export factor-1), which is known to be important for nuclear export of mRNA and proteins. Treatment with a TAP-specific siRNA strikingly shifted cytoplasmic HBc to nucleus, and led to a near 7-fold reduction of viral replication, and a near 10-fold reduction in HBsAg secretion. v) HBc of mutant ARD-II+IV was accumulated predominantly in the nucleus in a mouse model by hydrodynamic delivery. In addition to the revised map of NLS, our results suggest that HBc could shuttle rapidly between nucleus and cytoplasm via a novel TAP-dependent NES. |
format | Text |
id | pubmed-2965763 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2010 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-29657632010-11-08 Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles Li, Hung-Cheng Huang, Er-Yi Su, Pei-Yi Wu, Szu-Yao Yang, Ching-Chun Lin, Young-Sun Chang, Wen-Chang Shih, Chiaho PLoS Pathog Research Article It remains unclear what determines the subcellular localization of hepatitis B virus (HBV) core protein (HBc) and particles. To address this fundamental issue, we have identified four distinct HBc localization signals in the arginine rich domain (ARD) of HBc, using immunofluorescence confocal microscopy and fractionation/Western blot analysis. ARD consists of four tight clustering arginine-rich subdomains. ARD-I and ARD-III are associated with two co-dependent nuclear localization signals (NLS), while ARD-II and ARD-IV behave like two independent nuclear export signals (NES). This conclusion is based on five independent lines of experimental evidence: i) Using an HBV replication system in hepatoma cells, we demonstrated in a double-blind manner that only the HBc of mutant ARD-II+IV, among a total of 15 ARD mutants, can predominantly localize to the nucleus. ii) These results were confirmed using a chimera reporter system by placing mutant or wild type HBc trafficking signals in the heterologous context of SV40 large T antigen (LT). iii) By a heterokaryon or homokaryon analysis, the fusion protein of SV40 LT-HBc ARD appeared to transport from nuclei of transfected donor cells to nuclei of recipient cells, suggesting the existence of an NES in HBc ARD. This putative NES is leptomycin B resistant. iv) We demonstrated by co-immunoprecipitation that HBc ARD can physically interact with a cellular factor TAP/NXF1 (Tip-associated protein/nuclear export factor-1), which is known to be important for nuclear export of mRNA and proteins. Treatment with a TAP-specific siRNA strikingly shifted cytoplasmic HBc to nucleus, and led to a near 7-fold reduction of viral replication, and a near 10-fold reduction in HBsAg secretion. v) HBc of mutant ARD-II+IV was accumulated predominantly in the nucleus in a mouse model by hydrodynamic delivery. In addition to the revised map of NLS, our results suggest that HBc could shuttle rapidly between nucleus and cytoplasm via a novel TAP-dependent NES. Public Library of Science 2010-10-28 /pmc/articles/PMC2965763/ /pubmed/21060813 http://dx.doi.org/10.1371/journal.ppat.1001162 Text en Li et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited. |
spellingShingle | Research Article Li, Hung-Cheng Huang, Er-Yi Su, Pei-Yi Wu, Szu-Yao Yang, Ching-Chun Lin, Young-Sun Chang, Wen-Chang Shih, Chiaho Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles |
title | Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles |
title_full | Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles |
title_fullStr | Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles |
title_full_unstemmed | Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles |
title_short | Nuclear Export and Import of Human Hepatitis B Virus Capsid Protein and Particles |
title_sort | nuclear export and import of human hepatitis b virus capsid protein and particles |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2965763/ https://www.ncbi.nlm.nih.gov/pubmed/21060813 http://dx.doi.org/10.1371/journal.ppat.1001162 |
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