Characterization of the SUMO-Binding Activity of the Myeloproliferative and Mental Retardation (MYM)-Type Zinc Fingers in ZNF261 and ZNF198

Abstract

SUMO-binding proteins interact with SUMO modified proteins to mediate a wide range of functional consequences. Here, we report the identification of a new SUMO-binding protein, ZNF261. Four human proteins including ZNF261, ZNF198, ZNF262, and ZNF258 contain a stretch of tandem zinc fingers called myeloproliferative and mental retardation (MYM)-type zinc fingers. We demonstrated that MYM-type zinc fingers from ZNF261 and ZNF198 are necessary and sufficient for SUMO-binding and that individual MYM-type zinc fingers function as SUMO-interacting motifs (SIMs). Our binding studies revealed that the MYM-type zinc fingers from ZNF261 and ZNF198 interact with the same surface on SUMO-2 recognized by the archetypal consensus SIM. We also present evidence that MYM-type zinc fingers in ZNF261 contain zinc, but that zinc is not required for SUMO-binding. Immunofluorescence microscopy studies using truncated fragments of ZNF198 revealed that MYM-type zinc fingers of ZNF198 are necessary for localization to PML-nuclear bodies (PML-NBs). In summary, our studies have identified and characterized the SUMO-binding activity of the MYM-type zinc fingers in ZNF261 and ZNF198.

Screen for multi-SUMO–binding proteins reveals a multi-SIM–binding mechanism for recruitment of the transcriptional regulator ZMYM2 to chromatin

“…Recently, ZMYM2 was shown to bind specifically to the SUMO-modified form of HDAC1, but this interaction was driven through its zinc finger region rather than through the functionally important SIMs we have identified here (35). Moreover, a recent paper demonstrated mono-SUMO and poly-SUMO chain binding activity for ZMYM2, and again this was attributed to sequences contained in the zinc finger region of the protein (30). These findings support the idea that this protein might have different ways of interaction with SUMOylated partner proteins, but future studies are required to determine the precise nature of the multi-SUMOylated species that ZMYM2 binds to in the cell.…”

“…Given the strong multi-SIM-mediated binding characteristics of ZMYM2, we focused on this further. A previous study identified the zinc finger motifs of ZMYM2 as necessary and sufficient for binding to poly-SUMO chains (30). Importantly, full-length ZMYM2 lacking SIM2, which is critically important for binding the multi-SUMO scaffold, still retained poly-SUMO chain binding activity (Fig.…”

“…We therefore focused on the N-terminal region of ZMYM2, which contains two functional canonical SIMs, and asked whether this was sufficient for multi-SUMO binding. This region has previously been shown to be required for colocalization with PML and copurification of SUMOylated proteins (30). Although ZMYM2(1-200) was able to efficiently bind directly to His-tagged COMP-SUMO3 in pulldown assays, it was unable to bind to either dimeric or tetrameric SUMO chains (Fig.…”

Identification of Posttranslational Modification-Dependent Protein Interactions Using Yeast Surface Displayed Human Proteome Libraries

“…Numerous interactions were uncovered, confirming the importance of C/EBPβ methylation status in modulating interactions with other proteins. Recently, Guzzo et al used protein microarrays containing ~4,000 human transcription factors to conduct SUMO-binding assays and discovered that MYM-type zinc fingers from the transcription factors ZNF261 and ZNF198 function as SIMs [34]. …”

RAP80, ubiquitin and SUMO in the DNA damage response

“…The protein ZMYM3/ZNF261 was recently shown to bind to RAP80, Abraxas, and Bre1 at DSB sites [59]. ZMYM3/ZNF261 is recruited to DSBs in a RAP80-dependent manner, but also interacts directly with dsDNA, histone H2A/H2AX, and SUMO-2 polymeric chains [59, 60]. Unlike other members of the BRCA1-A complex such as RAP80, Abraxas, and BRCC36, depletion of ZMYM3/ZNF261 reduces HR repair, suggesting that ZMYM3/ZNF261 promotes HR [59].…”