PXD045052-1

PXD045052 is an original dataset announced via ProteomeXchange.

Title	Regulation of eDHFR-tagged proteins with trimethoprim PROTACs
Description	Temporal control of proteins in cells and living animals is crucial to improving the understanding of protein function in the post-genomic era. In addition, technologies that offer such control for engineered proteins could be used in therapeutic applications. Since regulation of proteins at a genomic or transcriptional level can be irreversible or a slow process, these tools may not be useful in settings where rapid temporal control is required to achieve immediate knockdown effects. PRoteolysis-TArgeting Chimeras (PROTACs) have emerged as a strategy to achieve rapid, post-translational control of protein abundance via recruitment of an E3 ligase to the target protein of interest. Here, we developed several PROTAC molecules by covalently linking the antibiotic trimethoprim (TMP) to pomalidomide, a small molecule ligand of the E3 ligase Cereblon. These molecules induce degradation of various proteins of interest (POIs) genetically fused to E. coli dihydrofolate reductase (eDHFR), the molecular target of TMP. We demonstrate that various eDHFR-tagged proteins, from fluorescent proteins to transcription factors and membrane-associated proteins, can be downregulated to 95% of maximum expression with our lead PROTAC molecule 7c. The data suggest that TMP-based PROTACs induce maximal degradation of POIs at drug concentrations that minimally affect the expression of immunomodulatory imide drug (IMiD)-sensitive neosubstrates. Finally, we show the ability to achieve multiplexed regulation with another known degron-PROTAC pair, and the formidable strength of our system for reversible protein regulation in a rodent model of metastatic cancer. Altogether, TMP PROTACs are a robust approach for selective and reversible degradation of eDHFR-tagged protein and have a strong potential for translation to in vivo models as well as dual degradation strategies with existing technologies.
HostingRepository	PRIDE
AnnounceDate	2023-12-06
AnnouncementXML	Submission_2023-12-06_14:31:59.971.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Richard Lauman
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Q Exactive HF

Revision	Datetime	Status	ChangeLog Entry
0	2023-09-02 20:26:57	ID requested
⏵ 1	2023-12-06 14:32:00	announced

Dataset with its publication pending

submitter keyword: eDHFR, Degradation, PROTACs

Richard Lauman
contact affiliation	Department of Radiology Perelman School of Medicine at the University of Pennsylvania
contact email	mark.sellmyer@pennmedicine.upenn.edu
lab head
Richard Lauman
contact affiliation	Universtiy of Pennsylvania
contact email	richard.lauman@gmail.com
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD045052
     1. Label: PRIDE project
     2. Name: Regulation of eDHFR-tagged proteins with trimethoprim PROTACs