Abstract
The protein inhibitor of activated STAT1 (PIAS1) is an E3 SUMO ligase that plays important roles in various cellular pathways. Increasing evidence shows that PIAS1 is overexpressed in various human malignancies, including prostate and lung cancers. Here we used quantitative SUMO proteomics to identify potential substrates of PIAS1 in a system-wide manner. We identified 983 SUMO sites on 544 proteins, of which 62 proteins were assigned as putative PIAS1 substrates. In particular, vimentin (VIM), a type III intermediate filament protein involved in cytoskeleton organization and cell motility, was SUMOylated by PIAS1 at Lys-439 and Lys-445 residues. VIM SUMOylation was necessary for its dynamic disassembly and cells expressing a non-SUMOylatable VIM mutant showed a reduced level of migration. Our approach not only enables the identification of E3 SUMO ligase substrates but also yields valuable biological insights into the unsuspected role of PIAS1 and VIM SUMOylation on cell motility.
Highlights
The protein inhibitor of activated signal transducers and activators of transcription 1 (STAT1) (PIAS1) is an E3 small ubiquitin-like modifier (SUMO) ligase that plays important roles in various cellular pathways
We further examined the phenotypic effects of protein inhibitor of activated STAT1 (PIAS1) expression on cell migration using the wound-healing assay
We noted that PIAS1-mediated SUMOylation occurred primarily on solvent-exposed lysine residues, which was the case for the global SUMOylome. These results suggest that PIAS1 may not impart conformational changes to its substrate upon binding, as it does not promote SUMOylation on lysine residues that would otherwise be buried within the core of the substrate
Summary
The protein inhibitor of activated STAT1 (PIAS1) is an E3 SUMO ligase that plays important roles in various cellular pathways. Several structurally unrelated classes of proteins appear to act as E3 SUMO ligases in mammalian cells, such as the protein inhibitor of activated STAT (PIAS) family of proteins, Ranbinding protein 2, the polycomb group protein (Pc2), and topoisomerase I- and p53-binding protein (TOPORS)[7,8]. Five different domains or motifs on PIAS family proteins recognize distinct sequences or conformations on target proteins, unique DNA structures, or specific “bridging” molecules to mediate their various functions[10]. An example of this is the SAF-A/B, Acinus and PIAS (SAP) domain, which has a strong affinity towards A–T-rich DNA11 and binds to Matrix attachment regions DNA12, in addition to having an important role in substrate recognition[13].
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