The Nuclear Dot Protein Sp100, Characterization of Domains Necessary for Dimerization, Subcellular Localization, and Modification by Small Ubiquitin-like Modifiers*
- From the Heinrich-Pette-Institut für Experimentelle Virologie und Immunologie an der Universität Hamburg, Martinistraße 52, D-20251 Hamburg, Germany
Abstract
The Sp100 and promyelocytic leukemia proteins (PML) are constituents of nuclear domains, known as nuclear dots (NDs) or PML bodies, and are both covalently modified by the small ubiquitin-related protein SUMO-1. NDs play a role in autoimmunity, virus infections, and in the etiology of acute promyelocytic leukemia. To date, little is known about the function of the Sp100 protein. Here we analyzed Sp100 domains that determine its subcellular localization, dimerization, and SUMOylation. A functional nuclear localization signal and an ND-targeting region that coincides with an Sp100 homodimerization domain were mapped. Sequences similar to the Sp100 homodimerization/ND-targeting region occur in several other proteins and constitute a novel protein motif, termed HSR domain. The lysine residue of the Sp100 protein, to which SUMO-1 is covalently linked, was mapped within and may therefore modulate the previously described HP1 protein-binding site. A consensus sequence for SUMOylation of proteins in general is suggested. SUMOylation strictly depended on a functional nuclear localization signal but was not necessary for nuclear import or ND targeting. A three-dimensional structure of Sp100, which supports the mapping data and provides additional information on Sp100 structure/function relationships, was generated by computer modeling. Taken together, our studies indicate the existence of well defined Sp100 domains with functions in ND targeting, nuclear import, nuclear SUMOylation, and protein-protein interaction.
Footnotes
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↵* This work was supported by grants from the Deutsche Forschungsgemeinschaft and the Deutsche Krebshilfe.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ To whom correspondence and reprint requests should be addressed: Tel./Fax: 0049 40 48051221; E-mail: sternsdo{at}hpi.uni-hamburg.de.
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↵2 T. Sternsdorf, E. Puccetti, K. Jensen, D. Hoelzer, H. Will, O. G. Ottmann, and M. Ruthardt, submitted for publication.
- Abbreviations:
- PML
-
promyelocytic leukemia protein
- Ab
-
antibody
- HP1
-
heterochromatin protein 1
- mAb
-
monoclonal antibody
- ND
-
nuclear dot(s) (nuclear domain(s) containing PML and Sp100 proteins)
- RAR
-
retinoic acid receptor-α
- SUMO-1
-
small ubiquitin-related modifier
- EGFP
-
enhanced green fluorescent protein
- HSR
-
homogeneously staining region
- CAT
-
chloramphenicol acetyltransferase
- aa
-
amino acid
-
- Received December 1, 1998.
- Revision received February 17, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
