A proteomic screen for nucleolar SUMO targets shows SUMOylation modulates the function of Nop5/Nop58.
Posttranslational SUMO modification is an important mechanism of regulating protein function, especially in the cell nucleus. The nucleolus is the subnuclear organelle responsible for rRNA synthesis, processing, and assembly of the large and small ribosome subunits. Here, we have used SILAC-based quantitative proteomics to identify nucleolar SUMOylated proteins. This reveals a role for SUMOylation in the biogenesis and/or function of small nucleolar ribonucleoprotein complexes (snoRNPs) via the targeting of Nhp2 and Nop58. Using combined in vitro and in vivo approaches, both Nhp2 and Nop58 (also known as Nop5) are shown to be substrates for SUMOylation. Mutational analyses revealed the sites of modification on Nhp2 as K5, and on Nop58 as K467 and K497. Unlike Nop58 and Nhp2, the closely related Nop56 and 15.5K proteins appear not to be SUMO targets. SUMOylation is essential for high-affinity Nop58 binding to snoRNAs. This study provides direct evidence linking SUMO modification with snoRNP function.
Westman
Belinda J
BJ
Wellcome Trust Centre for Gene Regulation and Expression, College of Life Sciences, University of Dundee, Dundee DD15EH, UK.
Verheggen
Céline
C
Hutten
Saskia
S
Lam
Yun Wah
YW
Bertrand
Edouard
E
Lamond
Angus I
AI
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002466:Cell Nucleolus / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008239:Lysine; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D040901:Proteomics / Q000379:methods; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism; D020718:Ribonucleoproteins, Small Nucleolar / Q000235:genetics / Q000378:metabolism*; D025842:SUMO-1 Protein / Q000378:metabolism; D025841:Small Ubiquitin-Related Modifier Proteins / Q000378:metabolism*; D014162:Transfection; D014452:Ubiquitins / Q000378:metabolism
D002868:Chromosomal Proteins, Non-Histone; C416929:NHP2 protein, human; C486600:NOP56 protein, human; C499062:NOP58 protein, human; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D017411:Ribonucleoproteins, Small Nuclear; D020718:Ribonucleoproteins, Small Nucleolar; D025842:SUMO-1 Protein; C495501:SUMO1 protein, human; C117909:SUMO2 protein, human; C492189:SUMO3 protein, human; D025841:Small Ubiquitin-Related Modifier Proteins; C402291:Snu13 protein, human; D014452:Ubiquitins; C047458:fibrillarin; D008239:Lysine
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