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Human Molecular Genetics Advance Access originally published online on March 17, 2004
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Human Molecular Genetics, 2004, Vol. 13, No. 10 1041-1048
DOI: 10.1093/hmg/ddh118
Human Molecular Genetics, Vol. 13, No. 10 © Oxford University Press 2004; all rights reserved

A potential role for the nucleolus in L1 retrotransposition

John L. Goodier1,*, Eric M. Ostertag1,2, Kurt A. Engleka1, Maria C. Seleme1 and Haig H. Kazazian, Jr1

1Department of Genetics, University of Pennsylvania School of Medicine and 2Department of Pathology and Laboratory Medicine, Hospital of the University of Pennsylvania, Philadelphia, PA, USA

Received January 13, 2004; Accepted March 9, 2004

Determining the subcellular localization of the L1 ORF2 protein (ORF2p) has been impossible to date because of technical limitations in detecting either endogenous or overexpressed forms of the protein. Here we report visualization of the full-length ORF2p in cultured human cells following expression in a modified vaccinia virus/T7 RNA polymerase (MVA/T7RP) system. The MVA/T7RP system was used to ascertain subcellular localization of L1 ORF1p and ORF2p both as fusions with green fluorescent protein and by immunocytochemistry. Full-length ORF2p was predominantly cytoplasmic, while carboxy-terminal-deleted ORF2p localized additionally to the nucleolus. We mapped a functional nucleolar localization signal in ORF2p. ORF1p appeared in the cytoplasm with a speckled pattern and colocalized with ORF2p in nucleoli in a subset of cells. These findings help explain the presence of chimeras between L1s and small RNA gene sequences recently discovered in the human genome.

* To whom correspondence should be addressed at: Department of Genetics, University of Pennsylvania School of Medicine, Room 515 CRB, 415 Curie Blvd, Philadelphia, PA 19104, USA. Tel: +1 2158982080; Fax: +1 2155737760; Email: jgoodier{at}mail.med.upenn.edu


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