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Identification of SENP6 targets in HeLa cells by SENP6 SiRNA and MW shift analysis

ABSTRACT: The SUMO protease SENP6 disassembles SUMO chains and thus SENP6 depletion should lead to the extension of SUMO polymers on substrates. We have used a proteomic method to identify putative SENP6 substrates based on increased apparent molecular weight after SENP6 depletion. This method uses SDS-PAGE fractionation of 6His-SUMO2 purifications from cells and slicing of gels into multiple pieces before tryptic digestion and MS analysis. Gel slices are assigned an average Apparent MW based on molecular weight standards (run in a separate lane), and this information combined with protein intensities to give every protein an Apparent MW both in the presence of SENP6 and its ablation via SiRNA. The difference between the two values allows us to understand which proteins become more SUMOylated upon SENP6 knock-down.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Permanent Cell Line Cell

SUBMITTER: Mike Tatham

LAB HEAD: Ronald T. Hay

PROVIDER: PXD036743 | Pride | 2024-03-01

REPOSITORIES: Pride

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