PXD046660

PXD046660 is an original dataset announced via ProteomeXchange.

Title	SUZ12 immunoprecipitation followed by protein indentification via mass spectrometry (SUZ12 IP-MS) in mESCs of SUZ12 WT, delta exon 4, KO, and KO rescues with SUZ12-L and SUZ12-S
Description	Suz12 exon 4 encodes 23 amino acids (aa 129–152 in SUZ12-L) that partially overlap with the WD-binding domain 1 (WDB1, 110–145). We reasoned that exon 4 skipping might alter the structure of SUZ12 and, possibly, PRC2 composition. To explore this possibility, we generated ESCs that lack the Suz12 exon 4 via CRISPR-Cas9–induced deletion. In addition, to rule out any biases due to the expression levels and/or SUZ12 epitope masking, we generated Suz12 knockout (KO) ESCs (herein, KO) in which Suz12 expression was subsequently rescued by re-introducing either the Suz12-L or Suz12-S mouse isoform fused to a triple-Flag tag under the regulation of a CAG promoter (KO+L/S; Figures S2H–S2K). We performed SUZ12 immunoprecipitation coupled with mass spectrometry (IP-MS) in the WT and ∆ex4 clones to compare their interactomes and with a flag antibody in the KO and rescue cell lines. As expected, no peptides corresponding to exon 4 were retrieved in ∆ex4 samples, while the rest of the sequence displayed similar coverage. Comparison of interactors in the two conditions revealed that SUZ12 binding to AEBP2 and JARID2 was strongly reduced in ∆ex4 cells with respect to WT cells, whereas SUZ12 binding to most core components or to PRC2.1-specific factors was unchanged or only slightly increased . These observations were confirmed by SUZ12 IP followed by Western blot (WB). Flag IP-MS in rescue cells confirmed that, while the long isoform was able to correctly form comparable amounts of both PRC2.1 and PRC2.2 subtypes, interaction of the SUZ12-S with PRC2.2-specific factors was drastically reduced.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_06:33:51.838.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Niccolò Arecco
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	No PTMs are included in the dataset
Instrument	LTQ Orbitrap Velos Pro

Revision	Datetime	Status	ChangeLog Entry
0	2023-11-03 14:05:24	ID requested
1	2024-03-18 03:53:47	announced
⏵ 2	2024-10-22 06:33:52	announced	2024-10-22: Updated project metadata.

10.1016/j.molcel.2024.02.011;

Arecco N, Mocavini I, Blanco E, Ballar, é C, Libman E, Bonnal S, Irimia M, Di Croce L, Alternative splicing decouples local from global PRC2 activity. Mol Cell, 84(6):1049-1061.e8(2024) [pubmed]

submitter keyword: Flag-tag, Rescues, KO, IP, PRC2, Interactions,mESCs, ESCs, SUZ12, Polycomb

Niccolò Arecco
contact affiliation	Centre for Genomic Regulation (CRG), Barcelona, Spain
contact email	arecco.niccolo@gmail.com
lab head
Niccolò Arecco
contact affiliation	CRG
contact email	arecco.niccolo@gmail.com
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2024/03/PXD046660

PRIDE project URI

• PRIDE
  1. PXD046660
     1. Label: PRIDE project
     2. Name: SUZ12 immunoprecipitation followed by protein indentification via mass spectrometry (SUZ12 IP-MS) in mESCs of SUZ12 WT, delta exon 4, KO, and KO rescues with SUZ12-L and SUZ12-S