Cargando…

In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles

The NHPX protein is a nucleolar factor that binds directly to a conserved RNA target sequence found in nucleolar box C/D snoRNAs and in U4 snRNA. Using enhanced yellow fluorescent protein (EYFP)– and enhanced cyan fluorescent protein–NHPX fusions, we show here that NHPX is specifically accumulated i...

Descripción completa

Detalles Bibliográficos
Autores principales: Leung, Anthony K.L., Lamond, Angus I.
Formato: Texto
Lenguaje:English
Publicado: The Rockefeller University Press 2002
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2173864/
https://www.ncbi.nlm.nih.gov/pubmed/12011111
http://dx.doi.org/10.1083/jcb.200201120
_version_ 1782145264183672832
author Leung, Anthony K.L.
Lamond, Angus I.
author_facet Leung, Anthony K.L.
Lamond, Angus I.
author_sort Leung, Anthony K.L.
collection PubMed
description The NHPX protein is a nucleolar factor that binds directly to a conserved RNA target sequence found in nucleolar box C/D snoRNAs and in U4 snRNA. Using enhanced yellow fluorescent protein (EYFP)– and enhanced cyan fluorescent protein–NHPX fusions, we show here that NHPX is specifically accumulated in both nucleoli and Cajal bodies (CBs) in vivo. The fusion proteins display identical localization patterns and RNA binding specificities to the endogenous NHPX. Analysis of a HeLa cell line stably expressing EYFP–NHPX showed that the nucleolar accumulation of NHPX was preceded by its transient accumulation in splicing speckles. Only newly expressed NHPX accumulated in speckles, and the nucleolar pool of NHPX did not interchange with the pool in speckles, consistent with a unidirectional pathway. The transient accumulation of NHPX in speckles prior to nucleoli was observed in multiple cell lines, including primary cells that lack CBs. Inhibitor studies indicated that progression of newly expressed NHPX from speckles to nucleoli was dependent on RNA polymerase II transcription, but not on RNA polymerase I activity. The data show a specific temporal pathway involving the sequential and directed accumulation of NHPX in distinct subnuclear compartments, and define a novel mechanism for nucleolar localization.
format Text
id pubmed-2173864
institution National Center for Biotechnology Information
language English
publishDate 2002
publisher The Rockefeller University Press
record_format MEDLINE/PubMed
spelling pubmed-21738642008-05-01 In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles Leung, Anthony K.L. Lamond, Angus I. J Cell Biol Article The NHPX protein is a nucleolar factor that binds directly to a conserved RNA target sequence found in nucleolar box C/D snoRNAs and in U4 snRNA. Using enhanced yellow fluorescent protein (EYFP)– and enhanced cyan fluorescent protein–NHPX fusions, we show here that NHPX is specifically accumulated in both nucleoli and Cajal bodies (CBs) in vivo. The fusion proteins display identical localization patterns and RNA binding specificities to the endogenous NHPX. Analysis of a HeLa cell line stably expressing EYFP–NHPX showed that the nucleolar accumulation of NHPX was preceded by its transient accumulation in splicing speckles. Only newly expressed NHPX accumulated in speckles, and the nucleolar pool of NHPX did not interchange with the pool in speckles, consistent with a unidirectional pathway. The transient accumulation of NHPX in speckles prior to nucleoli was observed in multiple cell lines, including primary cells that lack CBs. Inhibitor studies indicated that progression of newly expressed NHPX from speckles to nucleoli was dependent on RNA polymerase II transcription, but not on RNA polymerase I activity. The data show a specific temporal pathway involving the sequential and directed accumulation of NHPX in distinct subnuclear compartments, and define a novel mechanism for nucleolar localization. The Rockefeller University Press 2002-05-13 /pmc/articles/PMC2173864/ /pubmed/12011111 http://dx.doi.org/10.1083/jcb.200201120 Text en Copyright © 2002, The Rockefeller University Press This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Article
Leung, Anthony K.L.
Lamond, Angus I.
In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles
title In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles
title_full In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles
title_fullStr In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles
title_full_unstemmed In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles
title_short In vivo analysis of NHPX reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles
title_sort in vivo analysis of nhpx reveals a novel nucleolar localization pathway involving a transient accumulation in splicing speckles
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2173864/
https://www.ncbi.nlm.nih.gov/pubmed/12011111
http://dx.doi.org/10.1083/jcb.200201120
work_keys_str_mv AT leunganthonykl invivoanalysisofnhpxrevealsanovelnucleolarlocalizationpathwayinvolvingatransientaccumulationinsplicingspeckles
AT lamondangusi invivoanalysisofnhpxrevealsanovelnucleolarlocalizationpathwayinvolvingatransientaccumulationinsplicingspeckles