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Molecular & Cellular Proteomics 5:2298-2310, 2006.
© 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

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Research

Distinct and Overlapping Sets of SUMO-1 and SUMO-2 Target Proteins Revealed by Quantitative Proteomics^*,S

Alfred C. O. Vertegaal^,, Jens S. Andersen^¶, Stephen C. Ogg^||, Ronald T. Hay^**, Matthias Mann and Angus I. Lamond^**,

From the Department of Molecular Cell Biology, Leiden University Medical Center, 2300 RC Leiden, The Netherlands, ^¶ Center for Experimental BioInformatics (CEBI), Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, DK-5230 Odense M, Denmark, ^|| Centre for Molecular Medicine, 61 Biopolis Drive (Proteos), Singapore 138673, Singapore, ^** Wellcome Trust Biocentre, University of Dundee, Dundee DD1 5EH, United Kingdom, and Department of Proteomics and Signal Transduction, Max Planck Institute of Biochemistry, Am Klopferspitz 18, D-82152 Martinsried, Germany

The small ubiquitin-like modifier (SUMO) family in vertebrates includes three different family members that are conjugated as post-translational modifications to target proteins. SUMO-2 and -3 are nearly identical but differ substantially from SUMO-1. We used quantitative proteomics to investigate the target protein preferences of SUMO-1 and SUMO-2. HeLa cells were established that stably express His₆-SUMO-1 or His₆-SUMO-2. These cell lines and control HeLa cells were labeled with stable arginine isotopes, and His₆-SUMOs were enriched from lysates using immobilized metal affinity chromatography. 53 SUMO-conjugated proteins were identified, including 44 novel SUMO targets. 25 proteins were preferentially conjugated to SUMO-1, 19 were preferentially conjugated to SUMO-2, and nine proteins were conjugated to both SUMO-1 and SUMO-2. SART1 was confirmed by immunoblotting to have both SUMO-1- and SUMO-2-linked forms at similar levels. SUMO-1 and SUMO-2 are thus shown to have distinct and overlapping sets of target proteins, indicating that SUMO-1 and SUMO-2 may have both redundant and non-redundant cellular functions. Interestingly, 14 of the 25 SUMO-1-conjugated proteins contain zinc fingers. Although both SUMO family members play roles in many cellular processes, our data show that sumoylation is strongly associated with transcription because nearly one-third of the identified target proteins are putative transcriptional regulators.

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