Dataset Information

Proteomics analysis with the WDR5 PROTACs

ABSTRACT: Active gene transcription requires accessible chromatin. Post-translational modifications of histone proteins modulate accessibility to target genes, a process that is controlled by multiple chromatin modifying enzymes, remodelers and epigenetic reader proteins. Histone H3K4 methylation serves as hallmark of actively transcribed genes and is introduced by histone methyltransferases (HMTs). For proper function of HMT activity, several adaptor proteins are required. One of these proteins is the WD-repeat containing protein 5 (WDR5) that acts as scaffolding component in HMT complexes and that has been associated with controlling transcription factors including MYC and long non-coding RNAs. The wide influence of dysfunctional HMTs complexes and the typically upregulated MYC levels in diverse tumor types has made WDR5 an attractive cancer drug target. Indeed, protein-protein interface inhibitors for two protein interaction interfaces on WDR5 have been developed. While such compounds only inhibit a subset of WDR5 interactions, chemically induced proteasomal degradation of WDR5 might be an elegant way to target all oncogenic function. In this study, we present the design, synthesis and evaluation of two diverse WDR5 degrader series based on two WIN site binding scaffolds. We show that linker nature and length are essential for successful degradation and strongly influences the degradation rate. In the presented datasets, we determined the intracellular degradation specificity of the WDR5 PROTACs (8g, 6, 17b, 14). We therefore treated MV4-11 cells with 8g and 17b, the corresponding ligands 6 and 14, or DMSO and quantified the induced degradation using a label free approach.

INSTRUMENT(S): Q Exactive HF-X

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Suspension Culture, Blood, Macrophage

DISEASE(S): Acute Leukemia

SUBMITTER: Nicola Berner

LAB HEAD: Prof. Dr. Bernhard Kuster

PROVIDER: PXD025257 | Pride | 2022-01-27

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	4465_RA1_P051358_S00_A00.raw	Raw
	4465_RA1_P051360_S00_A00.raw	Raw
	4465_RA1_P051361_S00_A00.raw	Raw
	4465_RA1_P051362_S00_A00.raw	Raw
	4465_RA1_P051363_S00_A00.raw	Raw

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Publications

Design, Synthesis, and Evaluation of WD-Repeat-Containing Protein 5 (WDR5) Degraders.

Dölle Anja A Adhikari Bikash B Krämer Andreas A Weckesser Janik J Berner Nicola N Berger Lena-Marie LM Diebold Mathias M Szewczyk Magdalena M MM Barsyte-Lovejoy Dalia D Arrowsmith Cheryl H CH Gebel Jakob J Löhr Frank F Dötsch Volker V Eilers Martin M Heinzlmeir Stephanie S Kuster Bernhard B Sotriffer Christoph C Wolf Elmar E Knapp Stefan S

Journal of medicinal chemistry 20210513 15

Histone H3K4 methylation serves as a post-translational hallmark of actively transcribed genes and is introduced by histone methyltransferase (HMT) and its regulatory scaffolding proteins. One of these is the WD-repeat-containing protein 5 (WDR5) that has also been associated with controlling long noncoding RNAs and transcription factors including MYC. The wide influence of dysfunctional HMT complexes and the typically upregulated MYC levels in diverse tumor types suggested WDR5 as an attractive ...[more]

PMID: 33980013

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Project description:Poly(ADP-ribose) Polymerase 1 (PARP-1) is a nuclear enzyme that catalyse the transfer of ADP-ribose units from NAD+ to several target proteins involved in cellular stress responses. Using the cell line WRL68, we have previously shown that the activation of PARP-1 induced by oxidative stress after H2O2 treatment lead to depletion of cellular NAD+ and ATP, which promote cell death. In this work, we used LC-MS/MS based phosphoproteomics to show that the oxidative damage induced by H2O2 treatment modified the phosphorylation of several proteins in WRL68 cells, most of them related to transcription process. Interestingly, genetic or pharmacological inhibition of PARP-1 reduced the phosphorylation of YAP1, a transcriptional coactivator involved in cell survival, in WRL68 cells after H2O2 treatment. Inhibition of PARP-1 increased cell viability after oxidative treatment and diminished the degradation of YAP1; the elimination of YAP1 by gene silencing abrogated the protective effect of PARP-1 inhibition, indicating that YAP1 is important for the survival of WRL68 cells after oxidative damage. In the same way, the supplementation of NAD+, substrate of PARP-1, on WRL68 cells suffering oxidative damage reduced the phosphorylation of YAP1, suggesting that the loss of cellular NAD+ caused by PARP-1 activation after oxidative treatment is responsible for the phosphorylation of YAP1. Finally, PARP-1 silencing after oxidative treatment diminished the activation of AMPK, a master regulator of cell metabolism implicated in the restoration of energy balance during metabolic stress. Since NAD+ supplementation reduced the phosphorylation of some AMPK substrates in WRL68 cells after oxidative treatment, we hypothesize that the loss of cellular NAD+ after PARP-1 activation may induce a metabolic stress that activates AMPK. In summary, we showed a new crucial role of PARP-1 in the response to oxidative stress in WRL68 cells; PARP-1 activation reduces the cell viability by promoting the phosphorylation and degradation of YAP1, which is mediated by the energy stress caused by PARP-1 activation.