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Journal of Proteome Research Jan 2023Accurate quantification of proteomics data is essential for revealing and understanding biological signaling processes. We have recently developed a chemical proteomic...
Accurate quantification of proteomics data is essential for revealing and understanding biological signaling processes. We have recently developed a chemical proteomic strategy termed phosphatase inhibitor beads and mass spectrometry (PIB-MS) to investigate endogenous phosphoprotein phosphatase (PPP) dephosphorylation signaling. Here, we compare the robustness and reproducibility of status quo quantification methods for optimal performance and ease of implementation. We then apply PIB-MS to an array of breast cancer cell lines to determine differences in PPP signaling between subtypes. Breast cancer, a leading cause of cancer death in women, consists of three main subtypes: estrogen receptor-positive (ER+), human epidermal growth factor receptor two positive (HER2+), and triple-negative (TNBC). Although there are effective treatment strategies for ER+ and HER2+ subtypes, tumors become resistant and progress. Furthermore, TNBC has few targeted therapies. Therefore, there is a need to identify new approaches for treating breast cancers. Using PIB-MS, we distinguished TNBC from non-TNBC based on subtype-specific PPP holoenzyme composition. In addition, we identified an increase in PPP interactions with Hippo pathway proteins in TNBC. These interactions suggest that phosphatases in TNBC play an inhibitory role on the Hippo pathway and correlate with increased expression of YAP/TAZ target genes both in TNBC cell lines and in TNBC patients.
Topics: Humans; Female; Triple Negative Breast Neoplasms; Proteomics; Reproducibility of Results; Signal Transduction; Phosphoprotein Phosphatases
PubMed: 36448918
DOI: 10.1021/acs.jproteome.2c00465 -
Chemistry (Weinheim An Der Bergstrasse,... Jun 2018Protein phosphatases, while long overlooked, have recently become appreciated as drivers of both normal- and disease-associated signaling events. As a result, the... (Review)
Review
Protein phosphatases, while long overlooked, have recently become appreciated as drivers of both normal- and disease-associated signaling events. As a result, the spotlight is now turning torwards this enzyme family and efforts geared towards the development of modern chemical tools for studying these enzymes are well underway. This Minireview focuses on the evolution of chemical activity probes, both optical and covalent, for the study of protein phosphatases. Small-molecule probes, global monitoring of phosphatase activity through the use of covalent modifiers, and targeted fluorescence-based activity probes are discussed. We conclude with an overview of open questions in the field and highlight the potential impact of chemical tools for studying protein phosphatases.
Topics: Animals; Biomarkers; Fluorescent Dyes; Humans; Kinetics; Phosphoprotein Phosphatases; Signal Transduction
PubMed: 29338103
DOI: 10.1002/chem.201705194 -
The FEBS Journal Feb 2021Protein phosphorylation is a major reversible post-translational modification. Protein phosphatases function as 'critical regulators' in signaling networks through... (Review)
Review
Protein phosphorylation is a major reversible post-translational modification. Protein phosphatases function as 'critical regulators' in signaling networks through dephosphorylation of proteins, which have been phosphorylated by protein kinases. A large understanding of their working has been sourced from animal systems rather than the plant or the prokaryotic systems. The eukaryotic protein phosphatases include phosphoprotein phosphatases (PPP), metallo-dependent protein phosphatases (PPM), protein tyrosine (Tyr) phosphatases (PTP), and aspartate (Asp)-dependent phosphatases. The PPP and PPM families are serine(Ser)/threonine(Thr)-specific phosphatases (STPs), while PTP family is Tyr specific. Dual-specificity phosphatases (DsPTPs/DSPs) dephosphorylate Ser, Thr, and Tyr residues. PTPs lack sequence homology with STPs, indicating a difference in catalytic mechanisms, while the PPP and PPM families share a similar structural fold indicating a common catalytic mechanism. The catalytic cysteine (Cys) residue in the conserved HCX R active site motif of the PTPs acts as a nucleophile during hydrolysis. The PPP members require metal ions, which coordinate the phosphate group of the substrate, followed by a nucleophilic attack by a water molecule and hydrolysis. The variable holoenzyme assembly of protein phosphatase(s) and the overlap with other post-translational modifications like acetylation and ubiquitination add to their complexity. Though their functional characterization is extensively reported in plants, the mechanistic nature of their action is still being explored by researchers. In this review, we exclusively overview the plant protein phosphatases with an emphasis on their mechanistic action as well as structural characteristics.
Topics: Biocatalysis; Catalytic Domain; Models, Molecular; Phosphoprotein Phosphatases; Phosphorylation; Plant Proteins; Protein Domains; Protein Subunits; Signal Transduction; Substrate Specificity
PubMed: 32542989
DOI: 10.1111/febs.15454 -
The Journal of Biological Chemistry Nov 2015A central theme in nervous system function is equilibrium: synaptic strengths wax and wane, neuronal firing rates adjust up and down, and neural circuits balance... (Review)
Review
A central theme in nervous system function is equilibrium: synaptic strengths wax and wane, neuronal firing rates adjust up and down, and neural circuits balance excitation with inhibition. This push/pull regulatory theme carries through to the molecular level at excitatory synapses, where protein function is controlled through phosphorylation and dephosphorylation by kinases and phosphatases. However, these opposing enzymatic activities are only part of the equation as scaffolding interactions and assembly of multi-protein complexes are further required for efficient, localized synaptic signaling. This review will focus on coordination of postsynaptic serine/threonine kinase and phosphatase signaling by scaffold proteins during synaptic plasticity.
Topics: Animals; Humans; Nerve Tissue Proteins; Neuronal Plasticity; Phosphoprotein Phosphatases; Phosphorylation; Protein Serine-Threonine Kinases; Synaptic Transmission
PubMed: 26453308
DOI: 10.1074/jbc.R115.657262 -
Cell Stress & Chaperones May 2020Protein phosphatase 5 (PP5) is a serine/threonine protein phosphatase that regulates many cellular functions including steroid hormone signaling, stress response,... (Review)
Review
Protein phosphatase 5 (PP5) is a serine/threonine protein phosphatase that regulates many cellular functions including steroid hormone signaling, stress response, proliferation, apoptosis, and DNA repair. PP5 is also a co-chaperone of the heat shock protein 90 molecular chaperone machinery that assists in regulation of cellular signaling pathways essential for cell survival and growth. PP5 plays a significant role in survival and propagation of multiple cancers, which makes it a promising target for cancer therapy. Though there are several naturally occurring PP5 inhibitors, none is specific for PP5. Here, we review the roles of PP5 in cancer progression and survival and discuss the unique features of the PP5 structure that differentiate it from other phosphoprotein phosphatase (PPP) family members and make it an attractive therapeutic target.
Topics: Breast Neoplasms; Catalytic Domain; Female; HSP90 Heat-Shock Proteins; Humans; Neoplasms; Nuclear Proteins; Phosphoprotein Phosphatases
PubMed: 32239474
DOI: 10.1007/s12192-020-01091-3 -
Frontiers in Bioscience (Landmark... Jan 2012Wip1 (PPM1D) is a stress responsive PP2C phosphatase that plays a key role in stress signaling. Although originally identified as a gene induced by p53 after genotoxic... (Review)
Review
Wip1 (PPM1D) is a stress responsive PP2C phosphatase that plays a key role in stress signaling. Although originally identified as a gene induced by p53 after genotoxic stress, we now know that Wip1 expression is additionally regulated by other mechanisms. Wip1 is not only a target of p53, but is also a target of other transcription factors, including Estrogen Receptor-alpha and NF-kappaB. Additionally, Wip1 expression is regulated by post-transcriptional mechanisms such as mRNA stabilization and alternative splicing. Upon induction, Wip1 dampens the stress response by dephosphorylating and inactivating proteins such as p53, p38 MAPK, and ATM, usually as part of a negative feedback loop. As a result, Wip1 functions to abrogate cell cycle checkpoints and inhibit senescence, apoptosis, DNA repair, and the production of inflammatory cytokines. Furthermore, Wip1 is overexpressed in several types of human cancers and has oncogenic functions. The regulation of Wip1, the role of Wip1 in stress signaling, and the cooperation of Wip1 with oncogenes in promoting tumorigenesis will be discussed in this review.
Topics: Gene Expression Regulation, Enzymologic; Humans; Phosphoprotein Phosphatases; Protein Phosphatase 2C; Stress, Physiological
PubMed: 22201816
DOI: 10.2741/3999 -
Biochemical Society Transactions Jun 2017Protein phosphatase-6 (PP6) is a member of the PPP family of Ser/Thr phosphatases involved in intracellular signaling. PP6 is conserved among all eukaryotes, and... (Review)
Review
Protein phosphatase-6 (PP6) is a member of the PPP family of Ser/Thr phosphatases involved in intracellular signaling. PP6 is conserved among all eukaryotes, and genetics in model organisms indicates it has non-redundant functions relative to other PPP phosphatases. PP6 functions in association with conserved SAPS subunits and, in vertebrate species, forms heterotrimers with Ankrd subunits. Multiple studies have demonstrated how PP6 exerts negative control at different steps of nuclear factor kappaB signaling. Expression of PP6 catalytic subunit and the PPP6R1 subunit is especially high in hematopoietic cells and lymphoid tissues. Recent efforts at conditionally knocking out genes for PP6c or PP6R1 (SAPS1) have revealed distinctive effects on development of and signaling in lymphocytes.
Topics: Animals; Gene Expression Regulation; Genes; Humans; Lymphocytes; NF-kappa B; Phosphoprotein Phosphatases; Protein Conformation; Signal Transduction
PubMed: 28620030
DOI: 10.1042/BST20160169 -
British Journal of Pharmacology Jun 2012The MMPs and their inhibitors [tissue inhibitor of MMPs (TIMPs)] form the mainstay of extracellular matrix homeostasis. They are expressed in response to numerous... (Review)
Review
The MMPs and their inhibitors [tissue inhibitor of MMPs (TIMPs)] form the mainstay of extracellular matrix homeostasis. They are expressed in response to numerous stimuli including cytokines and GPCR activation. This review highlights the importance of adrenoceptors and phosphoprotein phosphatases (PPP) in regulating MMPs in the cardiovascular system, which may help explain some of the beneficial effects of targeting the adrenoceptor system in tissue remodelling and will establish emerging crosstalk between these three systems. Although α- and β-adrenoceptor activation increases MMP but decreases TIMP expression, MMPs are implicated in the growth stimulatory effects of adrenoceptor activation through transactivation of epidermal growth factor receptor. Furthermore, they have recently been found to catalyse the proteolysis of β-adrenoceptors and modulate vascular tone. While the mechanisms underpinning these effects are not well defined, reversible protein phosphorylation by kinases and phosphatases may be key. In particular, PPP (Ser/Thr phosphatases) are not only critical in resensitization and internalization of adrenoceptors but also modulate MMP expression. The interrelationship is complex as isoprenaline (ISO) inhibits okadaic acid [phosphoprotein phosphatase type 1/phosphoprotein phosphatase type 2A (PP2A) inhibitor]-mediated MMP expression. While this may be simply due to its ability to transiently increase PP2A activity, there is evidence for MMP-9 that ISO prevents okadaic acid-mediated expression of MMP-9 through a β-arrestin, NF-κB-dependent pathway, which is abolished by knock-down of PP2A. It is essential that crosstalk between MMPs, adrenoceptors and PPP are investigated further as it will provide important insight into how adrenoceptors modulate cardiovascular remodelling, and may identify new targets for pharmacological manipulation of the MMP system.
Topics: Adrenergic Agonists; Adrenergic Antagonists; Animals; Enzyme Activation; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Humans; Matrix Metalloproteinase Inhibitors; Matrix Metalloproteinases; Molecular Targeted Therapy; Phosphoprotein Phosphatases; Phosphorylation; Protein Processing, Post-Translational; Protein Transport; Proteolysis; Receptors, Adrenergic; Signal Transduction
PubMed: 22364165
DOI: 10.1111/j.1476-5381.2012.01917.x -
Biochimica Et Biophysica Acta.... Jan 2019Normal functioning of the brain is dependent upon a complex web of communication between numerous cell types. Within neuronal networks, the faithful transmission of... (Review)
Review
Normal functioning of the brain is dependent upon a complex web of communication between numerous cell types. Within neuronal networks, the faithful transmission of information between neurons relies on an equally complex organization of inter- and intra-cellular signaling systems that act to modulate protein activity. In particular, post-translational modifications (PTMs) are responsible for regulating protein activity in response to neurochemical signaling. The key second messenger, cyclic adenosine 3',5'-monophosphate (cAMP), regulates one of the most ubiquitous and influential PTMs, phosphorylation. While cAMP is canonically viewed as regulating the addition of phosphate groups through its activation of cAMP-dependent protein kinases, it plays an equally critical role in regulating removal of phosphate through indirect control of protein phosphatase activity. This dichotomy of regulation by cAMP places it as one of the key regulators of protein activity in response to neuronal signal transduction throughout the brain. In this review we focus on the role of cAMP in regulation of the serine/threonine phosphatases protein phosphatase 1 (PP1) and protein phosphatase 2A (PP2A) and the relevance of control of PP1 and PP2A to regulation of brain function and behavior.
Topics: Animals; Brain; Cyclic AMP; Cyclic AMP Response Element-Binding Protein; Cyclic AMP-Dependent Protein Kinases; Dopamine and cAMP-Regulated Phosphoprotein 32; Enzyme Inhibitors; Humans; Intracellular Signaling Peptides and Proteins; Phosphoprotein Phosphatases; Phosphorylation; Protein Phosphatase 1; Protein Phosphatase 2; Signal Transduction
PubMed: 30401536
DOI: 10.1016/j.bbamcr.2018.09.006 -
International Journal of Molecular... Apr 2022The fact that overexpression of the yeast Ser/Thr protein phosphatase Ppz1 induces a dramatic halt in cell proliferation was known long ago, but only work in the last... (Review)
Review
The fact that overexpression of the yeast Ser/Thr protein phosphatase Ppz1 induces a dramatic halt in cell proliferation was known long ago, but only work in the last few years has provided insight into the molecular basis for this toxicity. Overexpression of Ppz1 causes abundant changes in gene expression and modifies the phosphorylation state of more than 150 proteins, including key signaling protein kinases such as Hog1 or Snf1. Diverse cellular processes are altered: halt in translation, failure to properly adapt to low glucose supply, acidification of the cytosol, or depletion of intracellular potassium content are a few examples. Therefore, the toxicity derived from an excess of Ppz1 appears to be multifactorial, the characteristic cell growth blockage thus arising from the combination of various altered processes. Notably, overexpression of the Ppz1 regulatory subunit Hal3 fully counteracts the toxic effects of the phosphatase, and this process involves intracellular relocation of the phosphatase to internal membranes.
Topics: Cell Cycle; Phosphoprotein Phosphatases; Phosphorylation; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 35457140
DOI: 10.3390/ijms23084304