Dataset Information

TMT-ABPP cysteine profiling of SETDB1 modulators in MCF7 native proteome

ABSTRACT: We prodiled the cystene engagement of SETDB1 modulators at 10 uM in native MCF7 proteome/lysate with or without spiked of recombinant SETDB1 proteins.

INSTRUMENT(S): Orbitrap Eclipse

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Ka Yang

LAB HEAD: Ka Yang

PROVIDER: PXD056190 | Pride | 2025-05-07

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	613211_ee05303_ky_mcf7_unc_13a16a_cys_f1_1.mzid.gz	Mzid
	613289_ee05304_ky_mcf7_unc_13a16a_cys_f1_2.mzid.gz	Mzid
	613290_ee05305_ky_mcf7_unc_13a16a_cys_f2_1.mzid.gz	Mzid
	613291_ee05306_ky_mcf7_unc_13a16a_cys_f2_2.mzid.gz	Mzid
	613292_ee05307_ky_mcf7_unc_13a16a_cys_f3_1.mzid.gz	Mzid

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Publications

Potent and selective SETDB1 covalent negative allosteric modulator reduces methyltransferase activity in cells.

Uguen Mélanie M Shell Devan J DJ Silva Madhushika M Deng Yu Y Li Fengling F Szewczyk Magdalena M MM Yang Ka K Zhao Yani Y Stashko Michael A MA Norris-Drouin Jacqueline L JL Waybright Jarod M JM Beldar Serap S Rectenwald Justin M JM Mordant Angie L AL Webb Thomas S TS Herring Laura E LE Arrowsmith Cheryl H CH Ackloo Suzanne S Gygi Steven P SP McGinty Robert K RK Barsyte-Lovejoy Dalia D Liu Pengda P Halabelian Levon L James Lindsey I LI Pearce Kenneth H KH Frye Stephen V SV

Nature communications 20250224 1

A promising drug target, SETDB1, is a dual methyl-lysine (Kme) reader and methyltransferase implicated in cancer and neurodegenerative disease progression. To help understand the role of the triple Tudor domain (3TD) of SETDB1, its Kme reader, we first identify a low micromolar potency small molecule ligand, UNC6535, which occupies simultaneously both the TD2 and TD3 reader binding sites. Further optimization leads to the discovery of UNC10013, a covalent 3TD ligand targeting Cys385 of SETDB1. U ...[more]

PMID: 39994194

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Project description:Posttranscriptional regulation is emerging as a key factor in glucocorticoid (GC)-mediated gene regulation. We investigated the role of the human glucocorticoid receptor (GR) as an RNA-binding protein and its effect on mRNA turnover in human airway epithelial cells. Cell treatment with the potent GC budesonide accelerated the decay of CCL2 mRNA (t1/2=8±1 min vs. 62±17 min in DMSO-treated cells) and CCL7 mRNA (t1/2=15±4 min vs. 114±37 min), but not that of CCL5 mRNA (t1/2=231±8 min vs. 266±5 min) in the BEAS-2B cell line. This effect was inhibited by pre-incubation with an anti-GR antibody, indicating that GR itself plays a role in the turnover of these transcripts. Co-immunoprecipitation and biotin pulldown experiments showed that GR associates with CCL2 and CCL7 mRNAs, but not CCL5 mRNA. These methods confirmed CCL2 mRNA targeting by GR in human primary airway epithelial cells. Association of the GR was localized to the 5’UTR of CCL2 mRNA, and further mapped to nucleotides 44-60. The collection of transcripts associated with GR, identified by immunoprecipitation of GR-mRNA complexes followed by microarray analysis, revealed 479 transcripts that associated with GR. Computational analysis of the primary sequence and secondary structures of these transcripts yielded a GC-rich motif, which was shown to bind to GR in vitro. This motif was used to predict binding of GR to an additional 7889 transcripts. These results indicate that cytoplasmic GR interacts with a subset of mRNA through specific sequences and can regulate turnover rates, suggesting a novel posttranscriptional role for GR as an RNA-binding protein. The BEAS-2B cell line is derived from human tracheal epithelium transformed by an adenovirus 12-SV40 hybrid virus. For isolation of glucocorticoid receptor-associated transcripts, cytoplasmic lysates obtained from 1.6x107 cells/condition, 4 replicates per condition, were used to generate a collection of transcripts associated with GR by immunoprecipitation of the GR-mRNA complexes. The RNA from these IP's were extracted and labeled with the Illumina TotalPrep RNA Amplification Kit which generates biotin-labeled cRNA. This was hybridized to Illumina's Sentrix HumanRef-8,v3 Expression BeadChips. The data from 3 replicates of GR-IP and 4 replicates of Control-IP were used for global analysis of GR-mRNA interactions.