PXD038050-3

PXD038050 is an original dataset announced via ProteomeXchange.

Title	Comparison of CX-4945 and SCG-CK2-1 as Inhibitors of CSNK2 using Quantitative Phosphoproteomics: Triple SILAC in combination with Inhibitor-Resistant CSNK2
Description	Specificity is a limiting factor when using small-molecule inhibitors to study protein kinase signalling. Since inhibitor-resistant kinase mutants (i.e., drug-resistant alleles) remain active in the presence of inhibitor, they facilitate validation of on-target effects. By combining an inhibitor-resistant kinase mutant with mass spectrometry-based phosphoproteomics, we previously devised a systematic strategy for reliable identification and validation of CSNK2 substrates. In this study, we use the same strategy to evaluate the selectivity of CX-4945, a clinical stage CSNK2 inhibitor, and SGC-CK2-1, a chemical probe selectively targeting CSNK2. Human osteosarcoma (U2OS) cells expressing exogenous wild-type CSNK2A1 (WT) or an inhibitor-resistant triple mutant (TM, V66A/H160D/I174A) were treated with CX-4945 or SGC-CK2-1 prior to analysis using triple SILAC (phospho)proteomics. The minority of phosphosites, 15% at 4 hours and 5% at 24 hours, that were significantly downregulated in response to CX-4945 treatment were determined to be CSNK2A1-dependent. By comparison, the majority of phosphosites, >55% at both 4 and 24 hours, that were significantly downregulated in response to SGC-CK2-1 were identified as CSNK2A1-dependent. This indicates that SGC-CK2-1 exhibits dramatically greater selectivity towards CSNK2A1 than CX-4945. Notably, utilization of SGC-CK2-1 in cells expressing TM-CSNK2A1 enabled the identification of 330 CSNK2A1-dependent phosphosites. Overall, this study highlights the utility of exploiting highly selective chemical probes together with inhibitor-resistant kinase mutants to identify bona fide kinase substrates.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_06:00:14.148.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD038050
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Kristina Jurcic
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	phosphorylated residue
Instrument	Q Exactive; LTQ Orbitrap Elite

Revision	Datetime	Status	ChangeLog Entry
0	2022-11-08 08:07:15	ID requested
1	2023-08-13 20:15:41	announced
2	2023-11-14 06:50:26	announced	2023-11-14: Updated project metadata.
⏵ 3	2024-10-22 06:00:14	announced	2024-10-22: Updated project metadata.

10.1016/J.CRCHBI.2023.100041;

10.6019/PXD038050;

submitter keyword: protein kinase CK2, SGC-CK2-1, U2OS cells, CSNK2, SILAC, CX-4945, phosphoproteomics, LC-MS/MS

David Litchfield
contact affiliation	Department of Biochemistry, Schulich School of Medicine & Dentistry, Western University Department of Oncology, Schulich School of Medicine & Dentistry, Western University, London, Ontario, Canada. N6A 5C1
contact email	litchfi@uwo.ca
lab head
Kristina Jurcic
contact affiliation	University of Western Ontario
contact email	kjurcic@uwo.ca
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD038050
     1. Label: PRIDE project
     2. Name: Comparison of CX-4945 and SCG-CK2-1 as Inhibitors of CSNK2 using Quantitative Phosphoproteomics: Triple SILAC in combination with Inhibitor-Resistant CSNK2