PXD069264-1

PXD069264 is an original dataset announced via ProteomeXchange.

Title	Dynamic translocation of Inside-Out proteins to the cell surface underlies cellular adaptation to cancer-induced stress.
Description	Inside-out (I-O) protein display, the non-canonical surface localization of intracellular proteins, represents an underexplored feature of tumor cell biology. Here, we map the molecular landscape and trafficking mechanisms that control the presentation of I-O proteins on cancer cell membranes. Employing APEX2-mediated proximity biotinylation and a custom antibody generation and validation platform, we identified approximately 140 high-confidence I-O proteins, primarily ribosomal, proteasomal, chaperone, and translation factors, notably enriched in protein families associated with stress-response pathways. Validation of 500 antibodies encompassing 40 I-O targets across seven tumor cell lines confirmed selective and robust surface localization, while in vivo imaging in mouse xenografts demonstrated pronounced and tumor-specific antibody accumulation. I-O proteins were absent on PBMCs and in normal tissues, indicating cancer cell selectivity. Functional analyses revealed that I-O protein tethering to the membrane is dependent on heparan sulfate interactions; enzymatic removal of these glycans led to the clearance of I-O proteins from the cell surface. Notably, the removed proteins returned to baseline levels within six hours, indicating a dynamic balance related to ER-Golgi trafficking and cellular stress. Nearly half of these I-O proteins overlapped with known stress granule components; however, stress elements that promote stress granule formation do not similarly affect surface display of I-O proteins. Furthermore, I-O proteins are present on standard cancer cell lines under lower stress levels needed to induce stress granule formation, suggesting parallel yet mechanistically distinct aspects of the stress response. These findings position I-O display as a new paradigm in protein trafficking, different from traditional secretion pathways and closely linked to stress response.
HostingRepository	PRIDE
AnnounceDate	2026-03-09
AnnouncementXML	Submission_2026-03-09_08:16:31.114.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Tomasz Slezak
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	acetylated residue; monohydroxylated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2025-10-09 15:43:23	ID requested
⏵ 1	2026-03-09 08:16:31	announced

Dataset with its publication pending

submitter keyword: Inside-Out proteins, Cell surface proteome,Tumor-specific biomarkers

Anthony A Kossiakoff
contact affiliation	Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL 60637 Institute for Biophysical Dynamics, The University of Chicago, Chicago, IL 60637.
contact email	koss@bsd.uchicago.edu
lab head
Tomasz Slezak
contact affiliation	University of Chicago
contact email	slezak@uchicago.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD069264
     1. Label: PRIDE project
     2. Name: Dynamic translocation of Inside-Out proteins to the cell surface underlies cellular adaptation to cancer-induced stress.