Dataset Information

System-Wide Identification of Wild-Type SUMO-2 Conjugation Sites

ABSTRACT: SUMOylation is a reversible post-translational modification regulating all nuclear processes. Identification of SUMOylation sites by mass spectrometry has been hampered by bulky tryptic fragments, which thus far necessitated the use of mutated SUMO. Here, we present a dataset generated through a SUMO-specific protease-based methodology which circumvents this problem, dubbed Protease-Reliant Identification of SUMO Modification (PRISM). PRISM allows for detection of SUMOylated proteins as well as identification of specific sites of SUMOylation while using wild-type SUMO. The method is generic and could be widely applied to study lysine post-translational modifications. PRISM was employed in combination with high-resolution mass spectrometry to identify SUMOylation sites from HeLa cells under standard growth conditions and in response to heat shock. 751 wild-type SUMOylation sites on endogenous proteins were identified, including 200 dynamic SUMO sites in response to heat shock. Thus, we have developed the first method capable of quantitatively studying wild-type mammalian SUMO at the site-specific and system-wide level.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell, Hela Cell

DISEASE(S): Cervix Carcinoma

SUBMITTER: Ivo Hendriks

LAB HEAD: Alfred C.O. Vertegaal

PROVIDER: PXD001798 | Pride | 2015-06-15

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	MaxQuantOutput.zip	Other
	QE_IA_PRISM_BR1_TR1.raw	Raw
	QE_IA_PRISM_BR1_TR2.raw	Raw
	QE_IA_PRISM_Heat_BR1_TR1.raw	Raw
	QE_IA_PRISM_Heat_BR1_TR2.raw	Raw

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Publications

System-wide identification of wild-type SUMO-2 conjugation sites.

Hendriks Ivo A IA D'Souza Rochelle C RC Chang Jer-Gung JG Mann Matthias M Vertegaal Alfred C O AC

Nature communications 20150615

SUMOylation is a reversible post-translational modification (PTM) regulating all nuclear processes. Identification of SUMOylation sites by mass spectrometry (MS) has been hampered by bulky tryptic fragments, which thus far necessitated the use of mutated SUMO. Here we present a SUMO-specific protease-based methodology which circumvents this problem, dubbed Protease-Reliant Identification of SUMO Modification (PRISM). PRISM allows for detection of SUMOylated proteins as well as identification of ...[more]

PMID: 26073453

Similar Datasets

Project description:Genotoxic agents can cause replication fork stalling in dividing cells due to DNA lesions, eventually leading to replication fork collapse when the damage is not repaired. Small Ubiquitin-like Modifiers (SUMOs) are known to counteract replication stress, nevertheless, only a small number of relevant SUMO target proteins are known. To address this, we have purified and identified SUMO-2 target proteins regulated by replication stress in human cells. The developed methodology enabled single step purification of His10-SUMO-2 conjugates under denaturing conditions with high yield and high purity. The methodology is generic and is widely applicable in the ubiquitin field. Following statistical analysis on five biological replicates, a total of 566 SUMO-2 targets were identified. After 2 hours of Hydroxyurea treatment, 10 proteins were up-regulated for SUMOylation and 2 proteins were down-regulated for SUMOylation, whereas after 24 hours, 35 proteins were up-regulated for SUMOylation and 13 proteins were down-regulated for SUMOylation. A site-specific approach was used to map over 1,000 SUMO-2 acceptor lysines in target proteins. A large subset of these identified proteins function in one network that consists of interacting replication factors, transcriptional regulators, DNA damage response factors including MDC1, ATR-interacting protein ATRIP, the Bloom syndrome protein and the BLM-binding partner RMI1, the crossover junction endonuclease EME1, BRCA1 and CHAF1A. Furthermore, centromeric proteins and signal transducers were dynamically regulated by SUMOylation upon replication stress. Our results uncover a comprehensive network of SUMO target proteins dealing with replication damage and provide a framework for detailed understanding of the role of SUMOylation to counteract replication stress. Ultimately, our study reveals how a post-translational modification is able to orchestrate a large variety of different proteins to integrate different nuclear processes with the aim of dealing with the induced DNA damage