Human Wrnip1 Is Localized in Replication Factories in a Ubiquitin-binding Zinc Finger-dependent Manner

doi:10.1074/jbc.M803219200

Human Wrnip1 Is Localized in Replication Factories in a Ubiquitin-binding Zinc Finger-dependent Manner^*

^{^‡}Institute of Biochemistry II and Cluster of Excellence “Macromolecular Complexes,” “J. W. Goethe” University, Theodor-Stern-Kai 7, D-60590 Frankfurt (Main), Germany, ^{^§}Molecular Carcinogenesis and Center for Biomedical Genetics, The Netherlands Cancer Institute, Plesmanlaan 121, 1066 CX Amsterdam, The Netherlands, ^{^¶}Center for Experimental Research and Medical Studies and Department of Biomedical Sciences and Human Oncology, University of Torino, Via Santena 7, 10126 Torino, Italy, and ^{^∥}Bioinformatics Group, Miltenyi Biotec GmbH, MACS Molecular Business Unit, 50829 Cologne, Germany

3 To whom correspondence should be addressed: Institute for Biochemistry II, Theodor-Stern-Kai 7, D-60590 Frankfurt am Main, Germany. E-mail: ivan.dikic{at}biochem2.de.

Abstract

Wrnip1 (Werner helicase-interacting protein 1) has been implicated in the bypass of stalled replication forks in bakers' yeast. However, the function(s) of human Wrnip1 has remained elusive so far. Here we report that Wrnip1 is distributed inside heterogeneous structures detectable in nondamaged cells throughout the cell cycle. In an attempt to characterize these structures, we found that Wrnip1 resides in DNA replication factories. Upon treatments that stall replication forks, such as UVC light, the amount of chromatin-bound Wrnip1 and the number of foci significantly increase, further implicating Wrnip1 in DNA replication. Interestingly, the nuclear pattern of Wrnip1 appears to extend to a broader landscape, as it can be detected in promyelocytic leukemia nuclear bodies. The presence of Wrnip1 into these heterogeneous subnuclear structures requires its ubiquitin-binding zinc finger (UBZ) domain, which is able to interact with different ubiquitin (Ub) signals, including mono-Ub and chains linked via lysine 48 and 63. Moreover, the oligomerization of Wrnip1 mediated by its C terminus is also important for proper subnuclear localization. Our study is the first to reveal the composite and regulated topography of Wrnip1 in the human nucleus, highlighting its potential role in replication and other nuclear transactions.

Footnotes

↵⁴ The abbreviations used are: PCNA, proliferating cell nuclear antigen; Ub, ubiquitin; UBZ, ubiquitin-binding zinc finger; PML, promyelocytic leukemia; GST, glutathione S-transferase; DMEM, Dulbecco's modified Eagle's medium; PBS, phosphate-buffered saline; SPR, surface plasmon resonance; MALLS, multiangle laser light scattering; PIPES, 1,4-piperazinediethanesulfonic acid; RPA, replication protein A; UBA, ubiquitin-associated domain; Hs, Homo sapiens.
↵⁵ N. Crosetto, M. Bienko, and I. Dikic, unpublished observations.
↵^* This work was supported by Deutsche Forschungsgemeineschaft Grant DI 931/3-1 and grants from the Cluster of Excellence “Macromolecular Complexes” of the Frankfurt “J. W. Goethe” University (to I. D.) and NWO-CW (to T. S.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵ The on-line version of this article (available at http://www.jbc.org) contains supplemental Figs. 1-3.
↵¹ Both authors equally contributed to this work.
↵² Recipient of the “J. Buchmann” scholarship.
- Received April 28, 2008.
- Revision received September 12, 2008.
The American Society for Biochemistry and Molecular Biology, Inc.