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Showing PRAG1 — PRAGMIN is a alias.

PRAG1

Inactive tyrosine-protein kinase PRAG1 · UniProt Q86YV5

Length
1406 aa
Mass
149.6 kDa
Annotated
2026-06-10
20 papers in source corpus 15 papers cited in narrative 15 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 7/7 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

PRAG1 (Pragmin/SgK223/PEAK2/NACK) is a catalytically inactive pseudokinase that operates as a multifunctional scaffold coordinating Rho-family GTPase signaling, Src-family kinase (SFK) regulation, and Notch-dependent transcription (PMID:16481321, PMID:21873224, PMID:25038227). It was first defined as a GTP-dependent effector of Rnd2 that stimulates RhoA/Rho-kinase signaling and cell contraction (PMID:16481321). PRAG1 is tyrosine-phosphorylated on its EPIYA motif (Y391/Y411/Y413) by SFKs and, in response to EGFR, then binds and sequesters the SH2 domain of C-terminal Src kinase (Csk); this both potentiates Csk activity in a feed-forward loop and sustains membrane-associated SFK activation, driving cell migration and invasion at focal adhesions (PMID:21873224, PMID:27116701, PMID:35740644, PMID:35059869). Despite lacking catalytic activity, PRAG1 induces cellular protein tyrosine phosphorylation by recruiting active kinases such as Csk and ABL, the latter promoting actin-rich filopodia and invasion (PMID:29503074, PMID:35740644). Structurally, flanking N- and C-terminal helices around the pseudokinase domain form a split helical dimerization (SHED) fold mediating high-affinity homodimerization and higher-order oligomerization, and also support heterotypic association with the related pseudokinase SgK269/PEAK1, which bridges PRAG1 to Grb2 and is required for JAK1/Stat3 activation (PMID:26215634, PMID:27531744, PMID:29079850, PMID:29503074). In a distinct nuclear role, PRAG1/NACK associates with the Notch ternary complex on DNA as a transcriptional coactivator, recruited via p300/CBP-acetylated Maml1 and required for RNA polymerase II recruitment and, together with the Integrator complex, RNAPII-S5P-dependent transcription of Notch targets; this function depends on ATP binding and hydrolysis, and PRAG1 is itself a Notch target gene forming a feed-forward loop essential for Notch-driven tumorigenesis and embryonic viability (PMID:25038227, PMID:28625977, PMID:34551776, PMID:36938545). PRAG1 additionally forms stress-induced phase-separated condensates via its αN/αJ helices that mediate cell contraction (PMID:40149915), and stabilizes SNAIL1 by binding FBXO11 to promote EMT (PMID:40743806).

Mechanistic history

Synthesis pass · year-by-year structured walk · 14 steps
  1. 2006 High

    Established the founding function of PRAG1 by placing it in a GTPase signaling axis, answering what upstream and downstream partners control its biological output.

    Evidence Yeast two-hybrid, GTP-dependent binding and RhoA activity assays, and siRNA knockdown with morphological readout in PC12/HeLa cells

    PMID:16481321

    Open questions at the time
    • Did not define the molecular mechanism linking Rnd2 binding to RhoA activation
    • No structural basis for the Rnd2 interaction
  2. 2011 High

    Defined how PRAG1 regulates SFK signaling, showing that EPIYA phosphorylation converts it into a Csk-sequestering scaffold that sustains SFK activity.

    Evidence Co-IP, subcellular fractionation, and tyrosine phosphorylation assays compared against H. pylori CagA

    PMID:21873224

    Open questions at the time
    • Did not resolve which SFK residues PRAG1 protects
    • Stoichiometry of cytoplasmic Csk sequestration unquantified
  3. 2014 High

    Revealed an unexpected nuclear function as a Notch transcriptional coactivator and demonstrated its requirement for tumorigenesis and development.

    Evidence ChIP, Co-IP, reporter assays, siRNA, mouse knockout, and xenograft tumorigenesis

    PMID:25038227

    Open questions at the time
    • Mechanism of recruitment to the Notch complex not yet defined
    • Unclear how a focal-adhesion scaffold reaches chromatin
  4. 2015 High

    Connected PRAG1 overexpression to JAK1/Stat3-driven invasion, identifying a cytokine-signaling output downstream of the scaffold.

    Evidence Retroviral overexpression, siRNA, Co-IP, luciferase reporter, kinase inhibitors, and transwell assays in pancreatic cells

    PMID:26215634

    Open questions at the time
    • Direct vs indirect nature of the SgK223-Stat3 association unresolved
    • How JAK1 is activated by a pseudokinase scaffold unknown
  5. 2016 High

    Showed PRAG1 functions within heterotypic PEAK-family complexes, establishing it as the functionally required node downstream of SgK269/PEAK1 for Stat3 and migration outputs.

    Evidence MS proteomics, Co-IP, pulldown, SEC, and CRISPR knockout with migration/Stat3 assays

    PMID:27531744

    Open questions at the time
    • Functional difference between homo- and heterotypic complexes not fully dissected
  6. 2016 High

    Clarified the Csk interaction as a reciprocal feed-forward loop in which Csk phosphorylates PRAG1 and PRAG1 potentiates Csk activity at focal adhesions.

    Evidence Co-IP, in vitro kinase assay, confocal co-localization, and phosphosite mutagenesis

    PMID:27116701

    Open questions at the time
    • Net cellular consequence of simultaneous Csk activation and sequestration not reconciled
  7. 2017 High

    Provided the structural basis for PRAG1 oligomerization, defining the SHED fold that drives homodimerization and PEAK1 heterotypic assembly.

    Evidence X-ray crystallography of the pseudokinase domain with flanking helices plus mutagenesis and oligomerization assays

    PMID:29079850

    Open questions at the time
    • Link between specific oligomeric states and signaling outputs not established
  8. 2017 High

    Defined the recruitment mechanism to the Notch complex, showing p300/CBP acetylation of Maml1 is required to bring NACK in for RNAPII recruitment.

    Evidence Co-IP, ChIP, reporter assays, acetylation-site mutagenesis, and p300/CBP inhibition

    PMID:28625977

    Open questions at the time
    • Why Maml2 fails to recruit NACK not explained at the structural level
  9. 2018 High

    Confirmed catalytic inactivity by structure while showing PRAG1 still drives tyrosine phosphorylation through Csk, and identified a dimerization residue (A1329) controlling Csk activation.

    Evidence X-ray crystallography of the C-terminus, A1329E mutagenesis, proteomics, and Csk activity assays

    PMID:29503074

    Open questions at the time
    • Full substrate repertoire of the Csk-PRAG1 axis incompletely mapped
  10. 2021 High

    Resolved the division of labor in Notch transcription, with NACK recruiting RNAPII and the Integrator complex driving its S5 phosphorylation.

    Evidence SEC, CSL-DNA affinity FPLC, LC-MS/MS, ChIP, siRNA, and HEK293T reconstitution

    PMID:34551776

    Open questions at the time
    • Direct physical contacts between NACK and RNAPII not mapped
    • Order of supercomplex assembly unresolved
  11. 2023 Medium

    Identified an ATP-dependent function gating PRAG1's coactivator role, with ATP binding required for Notch complex association and druggable by a small molecule.

    Evidence ATPase and nucleotide-binding assays, Co-IP with the NTC, inhibitor screen, and PDX models

    PMID:36938545

    Open questions at the time
    • Structural basis for ATP-dependent NTC binding not determined
    • Mechanism limited by abstract-level detail
  12. 2022 Medium

    Extended the SFK/Csk and ABL invasion mechanism to colorectal and pancreatic contexts, pinpointing Y413/Y411 as the SRC/EGFR-driven phosphosite required for transformation.

    Evidence Phosphoproteomics, mutagenesis (Y413), localization, ABL kinase assays, xenografts, co-culture, and phospho-blots

    PMID:35059869 PMID:35740644

    Open questions at the time
    • Y411 vs Y413 numbering/usage across studies not unified
    • Y411 single Co-IP without reciprocal validation in the secretome study
  13. 2025 Medium

    Demonstrated that PRAG1 forms stress-induced phase-separated condensates via αN/αJ helices that are functionally required for cell contraction, linking it to neuronal stress states.

    Evidence Live-cell condensate imaging, αN/αJ mutagenesis, stress induction, contraction readout, and iPSC-derived dopaminergic neurons

    PMID:40149915

    Open questions at the time
    • Relationship of condensates to the SHED-mediated oligomers unclear
    • Disease relevance in Parkinson's not causally established
  14. 2025 Medium

    Identified a new role in EMT through FBXO11 binding that stabilizes SNAIL1, connecting PRAG1 to ubiquitin-pathway regulation.

    Evidence Co-IP, overexpression/knockdown with EMT-marker Western blots, and proliferation assays in biliary epithelial cells

    PMID:40743806

    Open questions at the time
    • Single Co-IP without reciprocal validation
    • Whether PRAG1 directly competes with FBXO11 substrate binding unknown

Open questions

Synthesis pass · forward-looking unresolved questions
  • How PRAG1 partitions between its cytoplasmic scaffolding/SFK roles, nuclear Notch coactivator role, and condensate-forming state, and what signals govern this switching, remains unresolved.
  • No unified model linking ATP-dependence, oligomerization, and condensate formation
  • Spatial regulation between focal adhesions, chromatin, and condensates undefined

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0060090 molecular adaptor activity 3 GO:0140096 catalytic activity, acting on a protein 3 GO:0140110 transcription regulator activity 3 GO:0098772 molecular function regulator activity 2 GO:0140657 ATP-dependent activity 1
Localization
GO:0005634 nucleus 3 GO:0005829 cytosol 1 GO:0005856 cytoskeleton 1
Pathway
R-HSA-162582 Signal Transduction 3 R-HSA-74160 Gene expression (Transcription) 3 R-HSA-1266738 Developmental Biology 1
Partners
ABLCSKFBXO11GRB2MAML1RND2SGK269/PEAK1STAT3
Complex memberships
Notch transcriptional activation complexPEAK family (SgK223/SgK269) complex

Evidence

Reading pass · 15 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2006 Pragmin (PRAG1) was identified as a novel effector of Rnd2 GTPase; it binds specifically to GTP-loaded Rnd2 (but not other Rho family GTPases) in a GTP-dependent manner, and this interaction stimulates RhoA activity and induces cell contraction through the RhoA/Rho-kinase pathway. Knockdown of Pragmin by siRNA enhances neurite elongation in PC12 cells, placing Pragmin downstream of Rnd2 and upstream of RhoA. Yeast two-hybrid screen; in vivo and in vitro binding assays; RhoA activity assays; siRNA knockdown in PC12 cells; morphological readout in HeLa cells The Journal of biological chemistry High 16481321
2011 Pragmin/SgK223 is tyrosine-phosphorylated at its EPIYA motif by Src family kinases (SFKs), and phosphorylated Pragmin binds the SH2 domain of C-terminal Src kinase (Csk), sequestering Csk in the cytoplasm away from the membrane and preventing inactivation of membrane-associated SFKs. This establishes a positive feedback loop of SFK activation, because SFKs phosphorylate Pragmin, which then sequesters Csk. Co-immunoprecipitation; subcellular fractionation; tyrosine phosphorylation assays; functional reporter assays; comparison with H. pylori CagA EPIYA effector Proceedings of the National Academy of Sciences of the United States of America High 21873224
2015 SgK223/Pragmin overexpression in human pancreatic ductal epithelial cells promotes cell migration and invasion through enhanced JAK1 activation and subsequent Stat3 Tyr705 phosphorylation and transcriptional activity; SgK223 and Stat3 physically associate in vivo, and pharmacological inhibition of JAK or Stat3 blocks SgK223-driven motility and invasion. Retroviral overexpression; siRNA knockdown; immunoprecipitation; Western blot; luciferase reporter assay; selective kinase inhibitors; transwell migration/invasion assays Molecular cancer High 26215634
2016 Pragmin directly binds Csk via its tyrosine-phosphorylated EPIYA motif; this complex formation potentiates Csk kinase activity, and Csk in turn phosphorylates Pragmin on Y238, Y343, and Y391 (the EPIYA motif), creating a feed-forward Csk activation loop. Pragmin and Csk co-localize to focal adhesions, and their interaction induces elongated cell morphology and elevated cell scattering in a mutually dependent manner. Co-immunoprecipitation; in vitro kinase assay; confocal co-localization; cell morphology assays; mutational analysis of phosphorylation sites Cancer science High 27116701
2016 SgK223 and SgK269/PEAK1 form both homotypic and heterotypic complexes dependent on their CH (α-helical) and pseudokinase (PK) domains; SgK269 bridges SgK223 to Grb2, but SgK269 cannot activate Stat3 or efficiently enhance migration in SgK223 knockout cells, demonstrating that SgK223 is functionally required downstream of SgK269 for these outputs. Mass spectrometry-based proteomics; Co-immunoprecipitation; pulldown; size-exclusion chromatography; CRISPR/Cas9 knockout; cell migration assays; Stat3 activation assays The Journal of biological chemistry High 27531744
2017 Crystal structure of the SgK223 pseudokinase domain with flanking N- and C-terminal helices reveals that these helices engage in a novel fold (later termed SHED) to mediate high-affinity homodimerization, and additional regulatory interfaces on the pseudokinase domain support assembly of large open-ended oligomers. Both homo- and heterotypic association with SgK269 is mediated by the CH and PK regions. X-ray crystallography; structure-function mutagenesis; biochemical oligomerization assays Nature communications High 29079850
2018 Crystal structure of the Pragmin C-terminus (residues 906–1368) confirms a classical protein-kinase fold devoid of catalytic activity despite a conserved catalytic lysine (K997). Flanking N- and C-terminal extensions form an original dimerization domain; the A1329E mutation in the C-terminal extension destabilizes dimerization and reduces Csk activation. Pragmin uses the tyrosine kinase Csk to induce protein tyrosine phosphorylation in human cells, identified by proteomics. X-ray crystallography; site-directed mutagenesis (A1329E); mass spectrometry proteomics; in-cell tyrosine phosphorylation assays; Csk kinase activity assays Structure (London, England : 1993) High 29503074
2014 NACK (SgK223/PRAG1) associates with the Notch transcriptional activation complex on DNA, acts as a Notch transcriptional coactivator, and is required for Notch-mediated tumorigenesis. Homozygous NACK loss is embryonic lethal in mice. NACK is also a Notch target gene, establishing a feed-forward loop. Co-immunoprecipitation; chromatin immunoprecipitation (ChIP); luciferase reporter assays; siRNA knockdown; mouse genetic knockout; xenograft tumorigenesis assays Cancer research High 25038227
2017 p300/CBP acetylates Mastermind-like 1 (Maml1) at K188 and K189, and this acetylation is required to recruit NACK (SgK223) to the Notch1 ternary complex, which then leads to RNA polymerase II recruitment and transcriptional initiation. NACK is recruited by Maml1 and Maml3 but not Maml2. Co-immunoprecipitation; ChIP; luciferase reporter assays; site-directed mutagenesis of acetylation sites; p300/CBP inhibitor treatment Cancer research High 28625977
2021 NACK (SgK223) is required for recruitment of RNA polymerase II to Notch-dependent promoters, while the Integrator complex (INT) is required for RNAPII phosphorylation at serine 5 (RNAPII-S5P) to initiate transcription; both NACK and INT act coordinately as components of the Notch transcriptional supercomplex. Size exclusion chromatography; CSL-DNA affinity FPLC; LC-MS/MS; ChIP; siRNA knockdown; HEK293T transfection reconstitution assay; xenograft assays Cell communication and signaling : CCS High 34551776
2023 NACK (SgK223/PRAG1) binds and hydrolyzes ATP, and only ATP-bound NACK can bind to the Notch ternary complex (NTC); a small-molecule inhibitor (Z271-0326) targeting this ATP-dependent function blocks Notch-mediated transcription. ATPase activity assay; nucleotide-binding assay; co-immunoprecipitation with NTC; small-molecule inhibitor screen; PDX mouse models Molecular therapy oncolytics Medium 36938545
2022 PEAK2 (PRAG1/SgK223) localizes to focal adhesions of colorectal cancer cells, induces cellular protein tyrosine phosphorylation (despite catalytic inactivity), and activates ABL tyrosine kinase; ABL-dependent formation of actin-rich filopodia drives cell invasion. The main phosphorylation site Tyr413 (phosphorylated by SRC) is required for all these transforming activities. Phosphoproteomic analysis; siRNA knockdown; overexpression; confocal localization; actin cytoskeleton imaging; ABL kinase assays; site-directed mutagenesis (Y413); nude mouse xenograft Cancers Medium 35740644
2022 SGK223 (PRAG1) is phosphorylated at Y411 by c-Src and in response to EGFR; tyrosine-phosphorylated SGK223 at Y411 interacts with CSK, upregulating c-Src activity and promoting cell migration. hAMSC secretome suppresses this pathway to inhibit Panc1 cancer cell invasion. Co-culture system; Western blot for phospho-SGK223 Y411, phospho-Src Y416/Y530; co-immunoprecipitation; invasion assay Medical oncology (Northwood, London, England) Medium 35059869
2025 PRAG1 forms dynamic phase-separated condensates in cells mediated by its αN and αJ helices; these condensates are required for mediating cell contraction, as condensate-formation-deficient mutants lose this function. Spherical PRAG1 condensates form under diverse stress conditions and in dopaminergic neurons from a Parkinson's disease patient. Live-cell imaging of condensates; mutagenesis of αN and αJ helices; stress induction assays; cell contraction phenotypic readout; iPSC-derived dopaminergic neurons Biomolecules Medium 40149915
2025 PRAG1 binds to F-box protein FBXO11 and reverses FBXO11-mediated inhibition of SNAIL1 protein expression (i.e., PRAG1 prevents FBXO11-dependent degradation of SNAIL1), thereby promoting EMT and cell proliferation in biliary epithelial cells. Co-immunoprecipitation (PRAG1–FBXO11 interaction); overexpression/knockdown with Western blot for SNAIL1, EMT markers; cell proliferation assays Biochimica et biophysica acta. Molecular basis of disease Medium 40743806

Source papers

Stage 0 corpus · 20 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2006 Pragmin, a novel effector of Rnd2 GTPase, stimulates RhoA activity. The Journal of biological chemistry 70 16481321
2011 Mammalian Pragmin regulates Src family kinases via the Glu-Pro-Ile-Tyr-Ala (EPIYA) motif that is exploited by bacterial effectors. Proceedings of the National Academy of Sciences of the United States of America 53 21873224
2015 The pseudokinase SgK223 promotes invasion of pancreatic ductal epithelial cells through JAK1/Stat3 signaling. Molecular cancer 46 26215634
2017 Structure of SgK223 pseudokinase reveals novel mechanisms of homotypic and heterotypic association. Nature communications 39 29079850
2016 C-terminal Src kinase-mediated EPIYA phosphorylation of Pragmin creates a feed-forward C-terminal Src kinase activation loop that promotes cell motility. Cancer science 38 27116701
2018 Dimerization of the Pragmin Pseudo-Kinase Regulates Protein Tyrosine Phosphorylation. Structure (London, England : 1993) 33 29503074
2014 NACK is an integral component of the Notch transcriptional activation complex and is critical for development and tumorigenesis. Cancer research 29 25038227
2005 The rice mutant dwarf bamboo shoot 1: a leaky mutant of the NACK-type kinesin-like gene can initiate organ primordia but not organ development. Plant & cell physiology 28 16183700
2016 Homo- and Heterotypic Association Regulates Signaling by the SgK269/PEAK1 and SgK223 Pseudokinases. The Journal of biological chemistry 27 27531744
2017 Acetylation of Mastermind-like 1 by p300 Drives the Recruitment of NACK to Initiate Notch-Dependent Transcription. Cancer research 21 28625977
2015 NACK kinesin is required for metaphase chromosome alignment and cytokinesis in the moss Physcomitrella patens. Cell structure and function 20 25748359
2016 Silencing NACK by siRNA inhibits tumorigenesis in non-small cell lung cancer via targeting Notch1 signaling pathway. Oncology reports 17 26782286
2022 The inhibition of Panc1 cancer cells invasion by hAMSCs secretome through suppression of tyrosine phosphorylation of SGK223 (at Y411 site), c-Src (at Y416, Y530 sites), AKT activity, and JAK1/Stat3 signaling. Medical oncology (Northwood, London, England) 14 35059869
2017 The pseudokinases SgK269 and SgK223: A novel oncogenic alliance in human cancer. Cell adhesion & migration 12 29105536
2021 NACK and INTEGRATOR act coordinately to activate Notch-mediated transcription in tumorigenesis. Cell communication and signaling : CCS 10 34551776
2019 SHEDding light on the role of Pragmin pseudo-kinases in cancer. American journal of cancer research 9 30906642
2022 Oncogenic Signalling of PEAK2 Pseudokinase in Colon Cancer. Cancers 6 35740644
2025 PRAG1 promotes cholangiocyte epithelial-mesenchymal transition and liver fibrosis in biliary atresia. Biochimica et biophysica acta. Molecular basis of disease 4 40743806
2023 A novel chemical attack on Notch-mediated transcription by targeting the NACK ATPase. Molecular therapy oncolytics 2 36938545
2025 PRAG1 Condensation Drives Cell Contraction Under Stress. Biomolecules 1 40149915

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