Dataset Information

Identification of PWWP2A interactors using label-free quantitative affinity enrichment mass spectrometry

ABSTRACT: Chromatin structure and function is regulated by reader proteins recognizing histone modifications and/or histone variants. We recently identified PWWP2A as a binder for H2A.Z-containing nucleosomes (Pünzeler, S. et al., EMBO J 2017). Here, as part of a larger project trying to characterize PWWP2A, we performed pulldowns using PWWP2A as a bait protein. Label-free quantitative mass spectrometry-based proteomics was used to detect and relatively quantify interaction partners of PWWP2A.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Cervix Carcinoma

SUBMITTER: Mario Oroshi

LAB HEAD: Matthias Mann

PROVIDER: PXD010202 | Pride | 2018-08-30

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	20140929_QEp4_EvKe_SA_LC6_GFP_01.raw	Raw
	20140929_QEp4_EvKe_SA_LC6_GFP_02.raw	Raw
	20140929_QEp4_EvKe_SA_LC6_GFP_03.raw	Raw
	20140929_QEp4_EvKe_SA_LC6_PWWP2A_01.raw	Raw
	20140929_QEp4_EvKe_SA_LC6_PWWP2A_02.raw	Raw

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Publications

PWWP2A binds distinct chromatin moieties and interacts with an MTA1-specific core NuRD complex.

Link Stephanie S Spitzer Ramona M M RMM Sana Maryam M Torrado Mario M Völker-Albert Moritz C MC Keilhauer Eva C EC Burgold Thomas T Pünzeler Sebastian S Low Jason K K JKK Lindström Ida I Nist Andrea A Regnard Catherine C Stiewe Thorsten T Hendrich Brian B Imhof Axel A Mann Matthias M Mackay Joel P JP Bartkuhn Marek M Hake Sandra B SB

Nature communications 20181016 1

Chromatin structure and function is regulated by reader proteins recognizing histone modifications and/or histone variants. We recently identified that PWWP2A tightly binds to H2A.Z-containing nucleosomes and is involved in mitotic progression and cranial-facial development. Here, using in vitro assays, we show that distinct domains of PWWP2A mediate binding to free linker DNA as well as H3K36me3 nucleosomes. In vivo, PWWP2A strongly recognizes H2A.Z-containing regulatory regions and weakly bind ...[more]

PMID: 30327463

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Project description:Our group is interested in epithelial-to-mesenchymal transition (EMT), in particular, TGF-beta induced EMT. TGF-beta signalling has been shown to be an important factor in the induction of EMT and it has been demonstrated that adding TGF-beta to epithelial cells in culture is a convenient way to study the process of EMT. In response to TGF-beta, Smad2 and 3 are activated, and form complexes with Smad4, which then regulate transcription of target genes through interactions with other DNA binding transcription factors. In the induction of EMT, the activated Smads mediate transcriptional regulation through three families of transcription factors, resulting in repression of epithelial marker gene expression and activation of mesenchymal gene expression (Xu J, et al. 2009) Also investigated in this study is the role of H2A.Z in EMT. H2A.Z is an evolutionary conserved and a metazoan essential histone variant of the H2A class. Mice deficient in H2A.Z die during early development but the reason for this is unknown (Faast et al. 2001). Previously, our laboratory showed that the loss of H2A.Z in Xenpous laevis impaired cell movement required for the formation of the mesoderm and neural crest (Ridgway et al. 2004). Given that mesoderm formation is critically dependent upon EMT, we therefore wondered whether H2A.Z might be a chromatin regulator of EMT. We transfected MDCK cells with a lentiviral vector to express a construct encoding an shRNA targeting canine H2A.Z as we wanted to test the hypothesis that H2A.Z is involved in the maintenance of cellular identity and that its loss might trigger de-differentiation. In order to investigate changes in histone variant H2A.Z occupancy associated with TGF-beta induced epithelial-to-mesenchymal transition (EMT) we performed H2A.Z ChIP-Seq in untreated and TGFb-treated MDCK cells. The MDCK cell line has been extensively used as a model system for EMT because they convert fully from the epithelial to the mesenchymal state in response to TGF-beta. Please note that RNA-seq data generated in conjunction to this ChIP-seq data set were also deposited at ArrayExpress under accession number E-MTAB-5628 ( https://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-5628 ).