Dataset Information

TurboID-based proxisome mapping of the human glucocorticoid receptor in A549 cells

ABSTRACT: Exogenous glucocorticoids are frequently used to treat inflammatory disorders and as adjuncts in solid cancers. However, their use is associated with severe side effects and therapy resistance. Novel glucocorticoid receptor (GR) ligands with a patient-validated reduced side effect profile have not yet reached the clinic. GR is a member of the nuclear receptor family of transcription factors and heavily relies on interactions with coregulator proteins for its transcriptional activity. To elucidate the role of the GR interactome in the altered transcriptional activity of GR following treatment with agonists, antagonists or lead selective GR agonists and modulators (SEGRAMs), we generated comprehensive interactome maps by high-confidence proximity proteomics in lung epithelial carcinoma cells. We found that the GR antagonist RU486 and the SEGRAM Dagrocorat both reduced GR interaction with CREB-binding protein (CBP)/p300 and the mediator complex compared to the full agonist Dexamethasone. Our data offer new insights into the role differential coregulator protein recruitment in shaping specific GR-mediated transcriptional responses.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

SUBMITTER: An Staes

LAB HEAD: Karolien De Bosscher

PROVIDER: PXD042014 | Pride | 2024-05-24

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	H092177_1p_Ionopticks1_PM3_CMB-1095_KRGEV_LauraVM_1.raw	Raw
	H092179_1p_Ionopticks1_PM3_CMB-1095_KRGEV_LauraVM_5.raw	Raw
	H092181_1p_Ionopticks1_PM3_CMB-1095_KRGEV_LauraVM_9.raw	Raw
	H092183_1p_Ionopticks1_PM3_CMB-1095_KRGEV_LauraVM_13.raw	Raw
	H092185_1p_Ionopticks1_PM3_CMB-1095_KRGEV_LauraVM_17.raw	Raw

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Publications

Selective Modulation of the Human Glucocorticoid Receptor Compromises GR Chromatin Occupancy and Recruitment of p300/CBP and the Mediator Complex.

Van Moortel Laura L Verhee Annick A Thommis Jonathan J Houtman René R Melchers Diana D Delhaye Louis L Van Leene Chloé C Hellemans Madeleine M Gevaert Kris K Eyckerman Sven S De Bosscher Karolien K

Molecular & cellular proteomics : MCP 20240220 3

Exogenous glucocorticoids are frequently used to treat inflammatory disorders and as adjuncts for the treatment of solid cancers. However, their use is associated with severe side effects and therapy resistance. Novel glucocorticoid receptor (GR) ligands with a patient-validated reduced side effect profile have not yet reached the clinic. GR is a member of the nuclear receptor family of transcription factors and heavily relies on interactions with coregulator proteins for its transcriptional act ...[more]

PMID: 38387774

Similar Datasets

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.