Affinage

PPP1R14B

Protein phosphatase 1 regulatory subunit 14B · UniProt Q96C90

Length
147 aa
Mass
15.9 kDa
Annotated
2026-06-10
37 papers in source corpus 12 papers cited in narrative 12 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

PPP1R14B (PHI-1) is a phosphorylation-dependent inhibitor of protein phosphatase-1 (PP1) that couples upstream kinase signaling to PP1-controlled substrate phosphorylation and proteostasis (PMID:12144526, PMID:17022978). Phosphorylation at Thr57 — by ILK in vitro and by PKC in intact vascular smooth muscle — converts PHI-1 into an active inhibitor that binds the PP1 catalytic subunit at its active site, inhibiting myosin light-chain phosphatase and producing Ca2+-sensitized smooth-muscle contraction (PMID:12144526, PMID:16267107, PMID:17022978). Its inhibitory potency is encoded by the PHIN domain and is intrinsically weaker than that of its paralog CPI-17, accounting for its distinct functional behavior (PMID:32784920). In migrating cells PHI-1 concentrates at juxtamembrane foci and the trailing edge, where it is specifically required for retraction during migration (PMID:15083373, PMID:15522888). Through the PP1 axis, PHI-1 governs the stability and phosphorylation state of downstream substrates: it stabilizes Raf-1 protein to restrain ERK signaling (PMID:38136612), and in cancer contexts maintains phospho-STMN1 to limit microtubule stability and drive paclitaxel resistance (PMID:36484700), sustains RPS6KA1 to activate AKT/NF-κB signaling in hepatocellular carcinoma (PMID:39216602), and supports Wnt/β-catenin signaling via the AKT/GSK-3β axis (PMID:41950731). PHI-1's own abundance is controlled by ubiquitin-proteasomal turnover, antagonized by RPS27A-recruited USP9X-mediated deubiquitination (PMID:36484700).

Mechanistic history

Synthesis pass · year-by-year structured walk · 10 steps
  1. 2002 High

    Established that PHI-1 is an inhibitory phosphoprotein and identified Thr57 as the kinase-controlled switch that activates its inhibition of myosin light-chain phosphatase to sensitize contraction.

    Evidence In vitro ILK phosphorylation with Thr57Ala mutagenesis, MLCP activity assay, and demembranated smooth-muscle contraction assay

    PMID:12144526

    Open questions at the time
    • Did not establish which kinase phosphorylates Thr57 in intact cells
    • No structural detail of the PHI-1–PP1c interface
  2. 2004 Medium

    Defined the subcellular distribution of PHI-1, showing it occupies discrete juxtamembrane foci distinct from the diffuse CPI-17 pattern.

    Evidence Immunohistochemistry, confocal and electron microscopy, tissue fractionation in smooth muscle and embryonic tissues

    PMID:15083373

    Open questions at the time
    • Molecular basis of juxtamembrane targeting unknown
    • Functional consequence of focal localization not tested here
  3. 2004 Medium

    Linked PHI-1 to a non-muscle process by showing it is required for trailing-edge retraction during cell migration, connecting its localization to a specific motility step.

    Evidence siRNA knockdown, wound-healing and time-lapse migration assays, immunostaining in HeLa and endothelial cells

    PMID:15522888

    Open questions at the time
    • PP1 substrate mediating retraction not identified
    • Phosphorylation dependence of the migration role not tested
  4. 2005 Medium

    Identified the physiological kinase input, showing agonist-induced PHI-1 phosphorylation in vascular smooth muscle is PKC-dependent and pathway-distinct from ROCK-driven CPI-17 phosphorylation.

    Evidence Phospho-specific immunoblot with PKC/ROCK inhibitors and PMA in primary rat aortic VSMCs; pharmacological dissection in chicken smooth muscle

    PMID:16081075 PMID:16267107

    Open questions at the time
    • Relative contribution of PKC vs Rho-kinase varies by tissue and is unresolved
    • In vivo signaling context not addressed
  5. 2006 Medium

    Demonstrated the mechanism of inhibition is phosphorylation-enhanced direct binding to the PP1 active site, providing a biochemical model for force generation at constant Ca2+.

    Evidence Skinned fiber contractility, PHI-1–PP1c binding assay, and microcystin active-site competition

    PMID:17022978

    Open questions at the time
    • No atomic-resolution structure of the complex
    • Single-lab biochemistry
  6. 2020 Medium

    Mapped the determinant of inhibitory specificity to the PHIN domain, explaining why PHI-1 is a weak myosin phosphatase inhibitor relative to CPI-17.

    Evidence In vitro phosphatase inhibition, cellular myosin phosphorylation, chimeric/deletion constructs, and zebrafish knockdown-rescue

    PMID:32784920

    Open questions at the time
    • Does not explain what physiological substrates PHI-1 preferentially regulates given its weak MLCP activity
    • Residues conferring specificity within PHIN not pinpointed
  7. 2023 Medium

    Extended PHI-1's PP1-axis role beyond contraction to substrate proteostasis, showing it controls Raf-1 protein levels and, in cancer, phospho-STMN1 stability to govern microtubule dynamics and drug resistance, with its own abundance set by USP9X deubiquitination.

    Evidence siRNA/overexpression with tautomycin PP1 inhibition for Raf-1; reciprocal Co-IP, ubiquitination assay, phospho-mutant STMN1 rescue, and xenograft for STMN1

    PMID:36484700 PMID:38136612

    Open questions at the time
    • Direct PP1-dependent dephosphorylation events on these substrates not reconstituted
    • Mechanism by which PHI-1 selects substrates unclear
  8. 2024 Medium

    Implicated PHI-1 in hepatocellular carcinoma signaling by showing it maintains RPS6KA1 stability and phosphorylation to activate the AKT/NF-κB pathway.

    Evidence siRNA/overexpression, Co-IP, phospho-mutant RPS6KA1 rescue, and xenograft model

    PMID:39216602

    Open questions at the time
    • Whether RPS6KA1 regulation requires PP1 catalytic activity not directly tested
    • Single tumor context
  9. 2025 Low

    Connected PHI-1 to androgen receptor regulation via a JAK2/STAT3 positive feedback loop affecting enzalutamide sensitivity in prostate cancer.

    Evidence siRNA/overexpression, western blot of JAK2/STAT3/AR, and pharmacological STAT3 inhibition

    PMID:41067365

    Open questions at the time
    • Mechanism inferred from inhibitor and KD/OE without direct binding or epistasis
    • No demonstration of a direct PHI-1–STAT3 interaction
  10. 2026 Medium

    Validated PPP1R14B as a druggable target whose depletion suppresses Wnt/β-catenin signaling and remodels the tumor immune microenvironment, identifying a direct small-molecule binder.

    Evidence SPR and CETSA binding validation of Astragaloside IV, ubiquitination assay, ROS analysis, T-cell co-culture, and syngeneic anti-PD-1 model

    PMID:41950731

    Open questions at the time
    • Binding site on PPP1R14B not mapped
    • Whether immune effects require the PP1 inhibitory function not established

Open questions

Synthesis pass · forward-looking unresolved questions
  • How a single weak PP1 inhibitor selectively directs PP1 toward distinct substrates (myosin light chain, Raf-1, STMN1, RPS6KA1) across tissues and what governs its juxtamembrane targeting remain unresolved.
  • No structural model of PHI-1 substrate/PP1 targeting
  • Mechanism of cell-type-specific substrate selection unknown
  • In vivo physiological function of PHI-1 not defined by genetic loss-of-function

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0098772 molecular function regulator activity 3 GO:0140096 catalytic activity, acting on a protein 3
Localization
GO:0005886 plasma membrane 2
Pathway
R-HSA-162582 Signal Transduction 3 R-HSA-392499 Metabolism of proteins 2 R-HSA-397014 Muscle contraction 2
Partners
PPP1CAPPP1CCRAF1RPS27ARPS6KA1STMN1USP9X

Evidence

Reading pass · 12 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2002 ILK phosphorylates PHI-1 (PPP1R14B) at Thr57, the critical inhibitory site; Thr57Ala mutation eliminates phosphorylation by ILK; thiophosphorylated PHI-1 inhibits myosin light-chain phosphatase (MLCP) activity and induces Ca2+ sensitization of smooth-muscle contraction, whereas the Thr57Ala mutant does not. In-gel kinase assay, solution phosphorylation assay, phosphopeptide mapping, phospho-amino acid analysis, phospho-specific immunoblotting, site-directed mutagenesis, MLCP activity assay, demembranated smooth-muscle contraction assay The Biochemical journal High 12144526
2004 PHI-1 (PPP1R14B) localizes juxtamembrane in distinct foci along the plasma membrane of smooth muscle cells (distinct from diffuse CPI-17 distribution), as determined by confocal and electron microscopy; first detected in embryonic heart at E10 and later in smooth muscle and endothelial cells of adult tissues. Immunohistochemistry, confocal microscopy, electron microscopy, tissue fractionation Histochemistry and cell biology Medium 15083373
2004 Knockdown of PHI-1 (PPP1R14B) by siRNA reduced HeLa cell migration rate by ~45%, caused an elongated phenotype, and specifically impaired retraction of the trailing edge during migration without affecting protrusive activity; PHI-1 was concentrated at the trailing edge of migrating endothelial cells. siRNA knockdown, wound-healing migration assay, time-lapse microscopy, cell spreading/replating assay, immunostaining Journal of cell science Medium 15522888
2005 In intact vascular smooth muscle cells, agonist stimulation (angiotensin II, thrombin, U-46619) induces PHI-1 (PPP1R14B) phosphorylation; this phosphorylation is selectively mediated by PKC (blocked by GF-109203X or PKC downregulation, induced by PMA), distinct from ROCK-dependent CPI-17 phosphorylation. 2D-PAGE, immunoblot with phospho-specific PHI-1 antibody, pharmacological inhibitors (Y-27632, H-1152, GF-109203X), constitutively active RhoA expression, PMA stimulation in primary rat aortic VSMCs American journal of physiology. Cell physiology Medium 16267107
2005 In chicken smooth muscle, G-protein stimulation via Rho-kinase leads to PHI-1 (PPP1R14B) phosphorylation (inhibitable by both Y-27632 and GF-109203X), correlating with increased myosin light chain phosphorylation; PHI-1 appears to act as a functional CPI-17 analog in this tissue where CPI-17 is absent. Western blot, pharmacological inhibitors (Y-27632, GF109203X), GTPγS stimulation, myosin light chain phosphorylation assay FEBS letters Medium 16081075
2006 Phosphorylated PHI-1 (P-PHI-1), but not unphosphorylated PHI-1, increases MLC20 phosphorylation and force in avian smooth muscle at constant [Ca2+]; phosphorylation of PHI-1 increases its direct interaction with the PP1 catalytic subunit (PP1c), and microcystin reduces this binding, indicating PHI-1 inhibits myosin light chain phosphatase by interacting with the active site of PP1c. Skinned smooth muscle fiber contractility assay, MLC20 phosphorylation assay, binding assay (PHI-1 vs PP1c), microcystin competition FEBS letters Medium 17022978
2020 The PHIN (phosphatase holoenzyme inhibitory) domain, not the N- or C-termini, encodes the specificity difference between Cpi-17 (potent myosin phosphatase inhibitor) and Phi-1 (PPP1R14B; weak myosin phosphatase inhibitory activity); Phi-1 paralogs in zebrafish display weak myosin phosphatase inhibitory activity in vitro and do not alter myosin phosphorylation in cells, and cannot rescue Cpi-17 knockdown. In vitro phosphatase inhibition assay, cellular myosin phosphorylation assay, deletion and chimeric protein analysis, zebrafish knockdown rescue experiments International journal of molecular sciences Medium 32784920
2023 PPP1R14B is degraded via the ubiquitin-proteasome pathway; RPS27A recruits deubiquitinase USP9X to deubiquitinate and stabilize PPP1R14B. PPP1R14B in turn maintains phosphorylation and protein stability of STMN1 in a manner dependent on PP1 catalytic subunits α and γ, thereby decreasing α-tubulin acetylation and microtubule stability to promote paclitaxel resistance; rescue by wild-type but not phosphorylation-deficient STMN1 confirmed the mechanism. Co-IP, ubiquitination assay, siRNA knockdown, gain/loss-of-function, western blot, xenograft tumor model, microtubule stability assay, phosphorylation-deficient STMN1 rescue experiment Cancer research Medium 36484700
2023 PHI-1 (PPP1R14B) knockdown in HEK293 cells causes a 15-fold increase in ERK1/2 phosphorylation attributable to elevated Raf-1 protein levels (not Ras or B-Raf); PHI-1 knockdown stabilizes Raf-1 protein without affecting Raf-1 mRNA; ectopic PHI-1 overexpression suppresses Raf-1 protein in an inhibitory-potency-dependent manner; PP1 inhibition with tautomycin mimics PHI-1 function to reduce Raf-1, establishing a PHI-1–PP1 axis that governs Raf-1 proteostasis. siRNA knockdown, ectopic overexpression, western blot (ERK1/2, Raf-1, B-Raf, Ras), RT-PCR for Raf-1 mRNA, tautomycin PP1 inhibition assay, EGF stimulation/MEK pathway assay Biomolecules Medium 38136612
2024 PPP1R14B maintains the protein stability and phosphorylation of RPS6KA1 (RSK1) and positively regulates activation of the AKT/NF-κB pathway in hepatocellular carcinoma cells; rescue by wild-type but not phosphorylation-deficient RPS6KA1 confirmed the mechanism. siRNA knockdown, ectopic overexpression, western blot, Co-IP, phospho-mutant rescue experiment, xenograft model Biochimica et biophysica acta. Molecular cell research Medium 39216602
2025 PPP1R14B regulates androgen receptor (AR) expression through the JAK2/STAT3 pathway and forms a positive feedback loop with STAT3; PPP1R14B knockdown reduces AR levels and sensitizes prostate cancer cells to enzalutamide, while overexpression has the opposite effect. siRNA knockdown, ectopic overexpression, western blot (JAK2, STAT3, AR), cell proliferation/migration/invasion assays, pharmacological STAT3 inhibition Cellular signalling Low 41067365
2026 Astragaloside IV directly binds PPP1R14B (KD = 4.88 μM by SPR and CETSA), inducing its ubiquitin-proteasomal degradation; PPP1R14B depletion inhibits Wnt/β-catenin signaling via the AKT/GSK-3β axis and induces mitochondrial ROS, relieving CX3CL1 repression and promoting CD8+ T cell recruitment. Molecular docking, surface plasmon resonance (SPR), cellular thermal shift assay (CETSA), ubiquitination assay, ROS analysis, T cell co-culture, syngeneic mouse model with anti-PD-1 Phytomedicine Medium 41950731

Source papers

Stage 0 corpus · 37 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2002 Phosphorylation of the myosin phosphatase inhibitors, CPI-17 and PHI-1, by integrin-linked kinase. The Biochemical journal 117 12144526
2023 Protein Phosphatase 1 Subunit PPP1R14B Stabilizes STMN1 to Promote Progression and Paclitaxel Resistance in Triple-Negative Breast Cancer. Cancer research 66 36484700
2007 The Drosophila PNG kinase complex regulates the translation of cyclin B. Developmental cell 58 17199048
1997 Png-1, a nervous system-specific zinc finger gene, identifies regions containing postmitotic neurons during mammalian embryonic development. The Journal of comparative neurology 42 9130664
2010 The N-glycanase png-1 acts to limit axon branching during organ formation in Caenorhabditis elegans. The Journal of neuroscience : the official journal of the Society for Neuroscience 38 20130186
2002 Methionine synthase activity and sulphur amino acid levels in the rat liver tumour cells HTC and Phi-1. Biochemical pharmacology 30 11853689
1976 Studies on the role and mode of operation of the very-lysine-rich histones in eukaryote chromatin. Nuclear-magnetic-resonance studies on nucleoprotein and histone phi 1-DNA complexes from marine invertebrate sperm. European journal of biochemistry 23 985747
2018 Identification of PNG kinase substrates uncovers interactions with the translational repressor TRAL in the oocyte-to-embryo transition. eLife 22 29480805
2017 Control of PNG kinase, a key regulator of mRNA translation, is coupled to meiosis completion at egg activation. eLife 21 28555567
2021 LINC00466 Impacts Cell Proliferation, Metastasis and Sensitivity to Temozolomide of Glioma by Sponging miR-137 to Regulate PPP1R14B Expression. OncoTargets and therapy 19 33642868
2004 Migration and retraction of endothelial and epithelial cells require PHI-1, a specific protein-phosphatase-1 inhibitor protein. Journal of cell science 19 15522888
2005 Divergent kinase signaling mediates agonist-induced phosphorylation of phosphatase inhibitory proteins PHI-1 and CPI-17 in vascular smooth muscle cells. American journal of physiology. Cell physiology 17 16267107
1990 Amino acid sequences recognized by T cells: studies on a merozoite surface antigen from the FCQ-27/PNG isolate of Plasmodium falciparum. Immunology letters 17 1704345
2020 LncRNA PPP1R14B-AS1 Promotes Tumor Cell Proliferation and Migration via the Enhancement of Mitochondrial Respiration. Frontiers in genetics 13 33262783
2020 The Evolution of Duplicated Genes of the Cpi-17/Phi-1 (ppp1r14) Family of Protein Phosphatase 1 Inhibitors in Teleosts. International journal of molecular sciences 12 32784920
1980 Bacteriophage phi 1 as a gene-cloning vector in Bacillus subtilis. Molecular & general genetics : MGG 12 6258022
2024 PPP1R14B-mediated phosphorylation enhances protein stability of RPS6KA1 to promote hepatocellular carcinoma tumorigenesis. Biochimica et biophysica acta. Molecular cell research 11 39216602
2008 Reconstructing the origin of the Lapita Cultural Complex: mtDNA analyses of East Sepik Province, PNG. Journal of human genetics 11 18498001
2006 PHI-1 interacts with the catalytic subunit of myosin light chain phosphatase to produce a Ca(2+) independent increase in MLC(20) phosphorylation and force in avian smooth muscle. FEBS letters 11 17022978
2005 PHI-1 induced enhancement of myosin phosphorylation in chicken smooth muscle. FEBS letters 11 16081075
2022 In vitro assessment of roles of PPP1R14B in cervical and endometrial cancer. Tissue & cell 10 35679681
2004 Juxtamembrane localization of the protein phosphatase-1 inhibitor protein PHI-1 in smooth muscle cells. Histochemistry and cell biology 10 15083373
1981 Unusually infrequent cleavage with several endonucleases and physical map construction of Bacillus subtilis bacteriophage phi 1 DNA. Journal of virology 9 6262531
2010 Characterization of the temperature-sensitive mutations un-7 and png-1 in Neurospora crassa. PloS one 7 20502699
1980 Bacillus subtilis-phage phi 1 overcomes host-controlled restriction by producing BamNx inhibitor protein. Molecular & general genetics : MGG 7 6255284
2024 Mutations in nucleotide metabolism genes bypass proteasome defects in png-1/NGLY1-deficient Caenorhabditis elegans. PLoS biology 5 38991033
2022 Long noncoding RNA PPP1R14B-AS1 imitates microRNA-134-3p to facilitate breast cancer progression by upregulating LIM and SH3 protein 1. Oncology research 4 37303940
2021 The GNU subunit of PNG kinase, the developmental regulator of mRNA translation, binds BIC-C to localize to RNP granules. eLife 4 34250903
1975 Impaired humoral immunity in PNG Highlanders. Papua and New Guinea medical journal 4 1061441
2025 Rediscovery of PHI-1/PPP1R14B: Emerging Roles of Cellular PP1 Signaling Mediated by the PPP1R14B Gene Product in Multiple Cancers and Beyond. Biomolecules 3 40149880
1996 The J-coupling restrained molecular mechanics (JrMM) protocol--an efficient alternative for deriving DNA endocyclic torsion angle constraints. Part I: Correlation of endocyclic torsion angles and vicinal torsion angle phi 1'2'. Journal of biomolecular structure & dynamics 3 8723775
2023 PHI-1, an Endogenous Inhibitor Protein for Protein Phosphatase-1 and a Pan-Cancer Marker, Regulates Raf-1 Proteostasis. Biomolecules 2 38136612
1975 Viral mutation affecting bacteriophage phi 1 development in Bacillus subtilis 168. Journal of virology 2 804042
1998 A sequence highly similar to PNG is located on chromosome 22q12 in intron 15 of the LIMK-2 gene. Biochemistry and molecular biology international 1 9556220
2026 Astragaloside IV directly targets PPP1R14B to sensitize prostate cancer to anti-PD-1 immunotherapy by remodeling the CX3CL1/CD8+ T cell axis. Phytomedicine : international journal of phytotherapy and phytopharmacology 0 41950731
2025 PPP1R14B promotes the progression of prostate cancer by regulating the JAK2/STAT3/AR pathway and alters the sensitivity and resistance to enzalutamide. Cellular signalling 0 41067365
1983 [Transport of Rb+ via activated sodium channels of clone N 18 phi 1 neuroblastoma cells]. Ukrainskii biokhimicheskii zhurnal (1978) 0 6318414

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