PXD067415

PXD067415 is an original dataset announced via ProteomeXchange.

Title	Proteomics- and BRET-screens identify SPRY2 as interactor of active Ras the impacts on its membrane organisation
Description	K-Ras is regulated and active in nanoscale proteo-lipid domains of the plasma membrane. Few regulators of K-Ras membrane organisation have been identified so far. We here combined a primary TurboID-based proximal proteome screen with a secondary BRET-screen to identify nine novel bona fide interactors of the K-Ras G-domain that would modulate its membrane organisation. We focused our hit characterisation on one novel candidate, APLP2, and SPRY2, which was previously implicated as negative regulator of MAPK-signalling. While APLP2 appeared to indirectly bind to K-Ras via C-Raf, SPRY2 showed characteristics of a new effector protein. We show by co-immunoprecipitation and BRET-experiments that the C-terminal fragment of SPRY2 comprising residues 161-315 interacts more with K-RasG12V than the full-length protein. Both full length and C-terminal fragment localized to the membrane. SPRY2 plasma membrane localization was blocked if K-Ras membrane anchorage was inhibited. Likewise, binding to oncogenic K-Ras was disrupted by K-Ras-inhibitors. Mutations at the predicted interface of the K-Ras effector binding region and the C-terminal fragment of SPRY2 modulated the interaction. Our data further suggest that SPRY2 operates as homo- or hetero-di/oligomer with SPRY4. Both full length SPRY2 and its C-terminal fragment promote differentiation of muscle C2C12 cells, which requires inhibition of MAPK-signalling. We propose a model, wherein active K-Ras recruits SPRY-dimers via the C-terminus of SPRY2 to the plasma membrane where they bind directly to Ras and block access of effectors.
HostingRepository	PRIDE
AnnounceDate	2025-10-02
AnnouncementXML	Submission_2025-10-02_09:46:14.933.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Fiona Hood
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	biotinylated residue; acetylated residue; monohydroxylated residue; iodoacetamide derivatized residue
Instrument	Q Exactive HF; Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2025-08-16 08:29:20	ID requested
⏵ 1	2025-10-02 09:46:15	announced

Dataset with its publication pending

submitter keyword: TurboID,KRas, APLP2, SPRY2, BRET

Ian A Prior
contact affiliation	Department of Molecular and Clinical Cancer Medicine University of Liverpool United Kingdom
contact email	Iprior@liv.ac.uk
lab head
Fiona Hood
contact affiliation	University of Liverpool
contact email	fehood@liverpool.ac.uk
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD067415
     1. Label: PRIDE project
     2. Name: Proteomics- and BRET-screens identify SPRY2 as interactor of active Ras the impacts on its membrane organisation