PXD033488 is an
original dataset announced via ProteomeXchange.
Dataset Summary
Title | Human RIPK3 C-lobe phosphorylation is essential for necroptotic signaling |
Description | Necroptosis is a caspase-independent, pro-inflammatory mode of programmed cell death which relies on the activation of the terminal effector, MLKL, by the upstream protein kinase RIPK3. To mediate necroptosis, RIPK3 must stably interact with, and phosphorylate the pseudokinase domain of MLKL. While the precise molecular cue that prompts RIPK3 to activate MLKL is incompletely understood, it is known that RIPK3 is highly regulated by phosphorylation. Here, we sought to identify phosphorylation sites of RIPK3 and dissect their regulatory functions. Phosphoproteomics identified 21 phosphorylation sites in HT29 cells overexpressing human RIPK3. By comparing cells expressing wild-type and kinase-inactive D142N RIPK3, autophosphorylation sites and substrates of other cellular kinases were distinguished. Of these 21 phosphosites, extensive mutational analyses identified only pT224 and pS227 as crucial, synergistic sites for stable interaction with MLKL to promote necroptosis, while the recently reported activation loop phosphorylation at S164/T165 negatively regulate the kinase activity of RIPK3. Despite being able to phosphorylate MLKL to a similar or higher extent than the wild-type RIPK3, mutation of T224, S227, and RHIM in RIPK3 attenuated or compromised their abilities to mediate necroptosis. This finding highlights the stable recruitment of human MLKL by RIPK3 to the necrosome as an essential checkpoint in necroptosis signaling, which is independent from and precedes the phosphorylation of MLKL. |
HostingRepository | PRIDE |
AnnounceDate | 2023-11-14 |
AnnouncementXML | Submission_2023-11-14_08:56:05.943.xml |
DigitalObjectIdentifier | |
ReviewLevel | Peer-reviewed dataset |
DatasetOrigin | Original dataset |
RepositorySupport | Unsupported dataset by repository |
PrimarySubmitter | Jarrod Sandow |
SpeciesList | scientific name: Homo sapiens (Human); NCBI TaxID: 9606; |
ModificationList | phosphorylated residue |
Instrument | timsTOF Pro |
Dataset History
Revision | Datetime | Status | ChangeLog Entry |
0 | 2022-04-28 00:25:42 | ID requested | |
1 | 2022-07-30 01:31:56 | announced | |
⏵ 2 | 2023-11-14 08:56:06 | announced | 2023-11-14: Updated project metadata. |
Publication List
Dataset with its publication pending |
Keyword List
submitter keyword: TNF, programmed necrosis,Pseudokinase, protein kinase, RIPK1 |
Contact List
James Murphy |
contact affiliation | Division Head, WEHI |
contact email | jamesm@wehi.edu.au |
lab head | |
Jarrod Sandow |
contact affiliation | WEHI |
contact email | sandow@wehi.edu.au |
dataset submitter | |
Full Dataset Link List
Dataset FTP location
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PRIDE project URI |
Repository Record List
[ + ]
[ - ]
- PRIDE
- PXD033488
- Label: PRIDE project
- Name: Human RIPK3 C-lobe phosphorylation is essential for necroptotic signaling