The luciferase-based in vivo protein-protein interaction assay revealed that CHK1 promotes PP2A and PME-1 interaction.

Protein phosphatase 2A (PP2A) is an essential serine/threonine protein phosphatase, and its dysfunction is involved in the onset of cancer and neurodegenerative disorders. PP2A functions as a trimeric holoenzyme whose composition is regulated by the methyl-esterification (methylation) of the PP2A catalytic subunit (PP2Ac). Protein phosphatase methylesterase-1 (PME-1) is the sole PP2Ac methylesterase, and the higher PME-1 expression is observed in various cancer and neurodegenerative diseases. Apart from serving as a methylesterase, PME-1 acts as a PP2A inhibitory protein, binding directly to PP2Ac and suppressing its activity. The intricate function of PME-1 hinders drug development by targeting the PME-1/PP2Ac axis. This study applied the NanoBiT system, a bioluminescence-based protein interaction assay, to elucidate the molecular mechanism that modulates unknown PME-1/PP2Ac protein-protein interaction (PPI). Compound screening identified that the CHK1 inhibitors inhibited PME-1/PP2Ac association without affecting PP2Ac methylation levels. CHK1 directly phosphorylates PP2Ac to promote PME-1 association. Phospho-mass spectrometry identified multiple phospho-sites on PP2Ac, including the Thr219, that affect PME-1 interaction. An anti-phospho-Thr219 PP2Ac antibody was generated and showed that CHK1 regulates the phosphorylation levels of this site in cells. On the contrary, in vitro phosphatase assay showed that CHK1 is the substrate of PP2A, and PME-1 hindered PP2A-mediated dephosphorylation of CHK1. Our data provides novel insights into the molecular mechanisms governing the PME-1/PP2Ac PPI and the triad relationship between PP2A, PME-1, and CHK1.

Transcriptome analysis revealed that PME-1 suppresses inflammatory signaling, activates PI3K/Akt signaling, and promotes epithelial-mesenchymal transition.

Protein phosphatase 2A (PP2A) is an essential serine/threonine protein phosphatase that belongs to the type2A protein phosphatase family with PP4 and PP6. PP2A functions as a trimeric holoenzyme, and the composition of the trimer is regulated by the methyl-esterification (methylation) of PP2A. Demethylation of PP2A is catalyzed by protein phosphatase methyl-esterase-1 (PME-1). Despite the physiological and pathophysiological importance of PME-1, the impact of changes in PME-1 expression on the transcriptome has not been reported. This study provides transcriptome data to gain a comprehensive understanding of the effects of PME-1 knockout on intracellular signaling of mouse embryonic fibroblasts. Our data showed that PME-1 suppresses inflammatory signaling, activates PI3K/Akt signaling, and promotes epithelial-mesenchymal transition.

Age-related alterations in protein phosphatase 2A methylation levels in brains of cynomolgus monkeys: a pilot study.

The abnormal activity of PP2A, a dominant member of type 2A serine/threonine protein phosphatase, has been implicated in the development of Alzheimer's disease (AD) and dementia with Lewy bodies (DLB). PP2A is a holoenzyme, and protein methylation of the catalytic subunit, PP2Ac, alters the complex composition. A decrease in PP2Ac methylation levels has been reported in AD and DLB. Aging is the most common risk factor for AD and DLB, but the relationship between aging and PP2A has not been studied in detail. Cynomolgus monkey show increased phosphorylation levels of tau and α-synuclein with aging. In this study, we investigated the alterations in the PP2A activity regulation with aging in monkey brains from 2 to 43 years of age using fractionated proteins. We found that type 2A protein phosphatase activity decreased with aging in cytoplasmic and nuclear-soluble fractions. PP2Ac methylation level was decreased in cytoplasmic and sarkosyl-insoluble fractions. A principal component analysis using PP2Ac, demethylated PP2Ac and PP2A methylesterase PME-1 levels in cytoplasmic and nuclear-soluble fractions as attributes showed that aged monkeys were in the same cluster. Our results show that brain aging in cynomolgus monkeys is closely related to changes in PP2A methylation.

Targets for Renal Carcinoma Growth Control Identified by Screening FOXD1 Cell Proliferation Pathways.

Clinical association studies suggest that FOXD1 is a determinant of patient outcome in clear cell renal cell carcinoma (ccRCC), and laboratory investigations have defined a role for this transcription factor in controlling the growth of tumors through regulation of the G2/M cell cycle transition. We hypothesized that the identification of pathways downstream of FOXD1 may define candidates for pharmacological modulation to suppress the G2/M transition in ccRCC. We developed an analysis pipeline that utilizes RNA sequencing, transcription factor binding site analysis, and phenotype validation to identify candidate effectors downstream from FOXD1. Compounds that modulate candidate pathways were tested for their ability to cause growth delay at G2/M. Three targets were identified: FOXM1, PME1, and TMEM167A, which were targeted by compounds FDI-6, AMZ-30, and silibinin, respectively. A 3D ccRCC tumor replica model was used to investigate the effects of these compounds on the growth of primary cells from five patients. While silibinin reduced 3D growth in a subset of tumor replicas, FDI-6 reduced growth in all. This study identifies tractable pathways to target G2/M transition and inhibit ccRCC growth, demonstrates the applicability of these strategies across patient tumor replicas, and provides a platform for individualized patient testing of compounds that inhibit tumor growth.

Coupling to short linear motifs creates versatile PME-1 activities in PP2A holoenzyme demethylation and inhibition.

Protein phosphatase 2A (PP2A) holoenzymes target broad substrates by recognizing short motifs via regulatory subunits. PP2A methylesterase 1 (PME-1) is a cancer-promoting enzyme and undergoes methylesterase activation upon binding to the PP2A core enzyme. Here, we showed that PME-1 readily demethylates different families of PP2A holoenzymes and blocks substrate recognition in vitro. The high-resolution cryoelectron microscopy structure of a PP2A-B56 holoenzyme-PME-1 complex reveals that PME-1 disordered regions, including a substrate-mimicking motif, tether to the B56 regulatory subunit at remote sites. They occupy the holoenzyme substrate-binding groove and allow large structural shifts in both holoenzyme and PME-1 to enable multipartite contacts at structured cores to activate the methylesterase. B56 interface mutations selectively block PME-1 activity toward PP2A-B56 holoenzymes and affect the methylation of a fraction of total cellular PP2A. The B56 interface mutations allow us to uncover B56-specific PME-1 functions in p53 signaling. Our studies reveal multiple mechanisms of PME-1 in suppressing holoenzyme functions and versatile PME-1 activities derived from coupling substrate-mimicking motifs to dynamic structured cores.

Hyperphosphorylation of Tau Due to the Interference of Protein Phosphatase Methylesterase-1 Overexpression by MiR-125b-5p in Melatonin Receptor Knockout Mice.

Melatonin has been indicated to ameliorate tau hyperphosphorylation in the pathogenesis of tau diseases, but the role of melatonin-receptor signal transduction has not been clearly discovered. In this study, we found intensive tau hyperphosphorylation in melatonin receptor knockout mice. Bielschowsky silver staining showed ghostlike neurofibrillary tangles in melatonin receptor-2 knockout (MT2KO) as well as melatonin receptors-1 and -2 knockout (DKO) mice, and an argyrophilic substance was deposited in melatonin receptor-1 knockout (MT1KO) mice. Furthermore, we found significantly decreased activity of protein phosphatase 2A (PP2A) by Western blot and enzyme-linked immunosorbent assay (ELISA), which was partly due to the overexpression of protein phosphatase methylesterase-1 (PME-1), but not glycogen synthase kinase-3ß (GSK-3ß), cyclin-dependent kinase 5 (CDK5) or protein kinase B (Akt). Finally, we observed a significant increase in cyclic adenosine monophosphate (cAMP) and a decrease in miR-125b-5p levels in MT1KO, MT2KO and DKO mice. Using a luciferase reporter assay, we discovered that miR-125b-5p largely decreased the expression of firefly luciferase by interfering with the 3'UTR of PME-1. Furthermore, miR-125b-5p mimics significantly decreased the expression of PME-1, while miR-125b-5p inhibitor induced tau hyperphosphorylation. These results show that melatonin-receptor signal transduction plays an important role in tau hyperphosphorylation and tangle formation.

Monotherapy efficacy of blood-brain barrier permeable small molecule reactivators of protein phosphatase 2A in glioblastoma.

Glioblastoma is a fatal disease in which most targeted therapies have clinically failed. However, pharmacological reactivation of tumour suppressors has not been thoroughly studied as yet as a glioblastoma therapeutic strategy. Tumour suppressor protein phosphatase 2A is inhibited by non-genetic mechanisms in glioblastoma, and thus, it would be potentially amendable for therapeutic reactivation. Here, we demonstrate that small molecule activators of protein phosphatase 2A, NZ-8-061 and DBK-1154, effectively cross the in vitro model of blood-brain barrier, and in vivo partition to mouse brain tissue after oral dosing. In vitro, small molecule activators of protein phosphatase 2A exhibit robust cell-killing activity against five established glioblastoma cell lines, and nine patient-derived primary glioma cell lines. Collectively, these cell lines have heterogeneous genetic background, kinase inhibitor resistance profile and stemness properties; and they represent different clinical glioblastoma subtypes. Moreover, small molecule activators of protein phosphatase 2A were found to be superior to a range of kinase inhibitors in their capacity to kill patient-derived primary glioma cells. Oral dosing of either of the small molecule activators of protein phosphatase 2A significantly reduced growth of infiltrative intracranial glioblastoma tumours. DBK-1154, with both higher degree of brain/blood distribution, and more potent in vitro activity against all tested glioblastoma cell lines, also significantly increased survival of mice bearing orthotopic glioblastoma xenografts. In summary, this report presents a proof-of-principle data for blood-brain barrier-permeable tumour suppressor reactivation therapy for glioblastoma cells of heterogenous molecular background. These results also provide the first indications that protein phosphatase 2A reactivation might be able to challenge the current paradigm in glioblastoma therapies which has been strongly focused on targeting specific genetically altered cancer drivers with highly specific inhibitors. Based on demonstrated role for protein phosphatase 2A inhibition in glioblastoma cell drug resistance, small molecule activators of protein phosphatase 2A may prove to be beneficial in future glioblastoma combination therapies.

Expression Pattern and Prognostic Utility of PME-1 in Patients with Hepatocellular Carcinoma.

PURPOSE: Hepatocellular carcinoma (HCC) remains one of the most common malignancies. While there is lack of markers capable of predicting which patients are at risk of aggressive course of the disease. Although a few protein phosphatase methyl-esterase-1 (PME-1) tumor-promoting mechanisms have been reported, the role of PME-1 in cancer including HCC occurrence and progression remains to be elucidated. The aim of this study was to explore the expression pattern and relationship between PME-1 with the pathological parameters in patients with HCC. METHODS: PME-1 expression was assessed from HCC tissue chips via immunohistochemistry. Chi-square test was used to identify the association between PME-1 staining and clinicopathological variables of HCC patients. Kaplan-Meier analysis and Cox regression analysis were performed to draw survival curves and verify the independent prognostic factors of HCC patients, respectively. RESULTS: We found that PME-1 expression was significantly higher in HCC tumor tissues compared with non-tumor tissues (P < 0.001). Furthermore, high expression level of PME-1 was significantly associated with differentiation (P = 0.047), tumor number (P = 0.001), intrahepatic or extrahepatic metastasis (P = 0.018), and recurrence (P = 0.001). Kaplan-Meier analysis revealed that high expression level of PME-1 was associated with shorter survival (P < 0.001). Univariate analysis with Log-rank test revealed that PME-1 expression status was significantly correlated with overall survival (P < 0.001). Furthermore, multivariate models with Cox proportional hazards analysis showed that high expression of PME-1 was a statistically independent predictive factor of poor prognosis in HCC patients (hazard ratio, 3.429; 95% confidence interval, 1.369-8.589; P = 0.009). CONCLUSION: In conclusion, these findings indicated that PME-1 expression was associated with aggressive pathological features and worse oncological outcomes, suggesting its potential therapeutic value for patients with HCC.

Glycogen synthase kinase-3ß suppresses the expression of protein phosphatase methylesterase-1 through ß-catenin.

Protein phosphatase 2A (PP2A) is the major tau phosphatase. Its activity toward tau is regulated by the methylation of PP2A catalytic subunit (PP2Ac) at Leu309. Protein phosphatase methylesterase-1 (PME-1) demethylates PP2Ac and suppresses its activity. We previously found that glycogen synthase kinase-3ß (GSK-3ß) suppresses PME-1 expression. However, the underlying molecular mechanism is unknown. In the present study, we analyzed the promoter of PME-1 gene and found that human PME-1 promoter contains two lymphoid enhancer binding factor-1/T-cell factor (LEF1/TCF) cis-elements in which ß-catenin serves as a co-activator. ß-catenin acted on these two cis-elements and promoted PME-1 expression. GSK-3ß phosphorylated ß-catenin and suppressed its function in promoting PME-1 expression. Inhibition and activation of GSK-3ß by PI3K-AKT pathway promoted and suppressed, respectively, PME-1 expression in primary cultured neurons, SH-SY5Y cells and in the mouse brain. These findings suggest that GSK-3ß phosphorylates ß-catenin and suppresses its function on PME-1 expression, resulting in an increase of PP2Ac methylation.

Arpp19 Promotes Myc and Cip2a Expression and Associates with Patient Relapse in Acute Myeloid Leukemia.

Disease relapse from standard chemotherapy in acute myeloid leukemia (AML) is poorly understood. The importance of protein phosphatase 2A (PP2A) as an AML tumor suppressor is emerging. Therefore, here, we examined the potential role of endogenous PP2A inhibitor proteins as biomarkers predicting AML relapse in a standard patient population by using three independent patient materials: cohort1 (n = 80), cohort2 (n = 48) and The Cancer Genome Atlas Acute Myeloid Leukemia (TCGA LAML) dataset (n = 160). Out of the examined PP2A inhibitors (CIP2A, SET, PME1, ARPP19 and TIPRL), expression of ARPP19 mRNA was found to be independent of the current AML risk classification. Functionally, ARPP19 promoted AML cell viability and expression of oncoproteins MYC, CDK1, and CIP2A. Clinically, ARPP19 mRNA expression was significantly lower at diagnosis (p = 0.035) in patients whose disease did not relapse after standard chemotherapy. ARPP19 was an independent predictor for relapse both in univariable (p = 0.007) and in multivariable analyses (p = 0.0001) and gave additive information to EVI1 expression and risk group status (additive effect, p = 0.005). Low ARPP19 expression was also associated with better patient outcome in the TCGA LAML cohort (p = 0.019). In addition, in matched patient samples from diagnosis, remission and relapse phases, ARPP19 expression was associated with disease activity (p = 0.034), indicating its potential usefulness as a minimal residual disease (MRD) marker. Together, these data demonstrate the oncogenic function of ARPP19 in AML and its risk group independent role in predicting AML patient relapse tendency.

PP2A: A Promising Biomarker and Therapeutic Target in Endometrial Cancer.

Over the last decade, the use of targeted therapies has immensely increased in the treatment of cancer. However, treatment for endometrial carcinomas (ECs) has lagged behind, although potential molecular markers have been identified. This is particularly problematic for the type II ECs, since these aggressive tumors are usually not responsive toward the current standard therapies. Therefore, type II ECs are responsible for most EC-related deaths, indicating the need for new treatment options. Interestingly, molecular analyses of type II ECs have uncovered frequent genetic alterations (up to 40%) in PPP2R1A, encoding the Aα subunit of the tumor suppressive heterotrimeric protein phosphatase type 2A (PP2A). PPP2R1A mutations were also reported in type I ECs and other common gynecologic cancers, albeit at much lower frequencies (0-7%). Nevertheless, PP2A inactivation in the latter cancer types is common via other mechanisms, in particular by increased expression of Cancerous Inhibitor of PP2A (CIP2A) and PP2A Methylesterase-1 (PME-1) proteins. In this review, we discuss the therapeutic potential of direct and indirect PP2A targeting compounds, possibly in combination with other anti-cancer drugs, in EC. Furthermore, we investigate the potential of the PP2A status as a predictive and/or prognostic marker for type I and II ECs.

Targeted therapies don't work for a reason; the neglected tumor suppressor phosphatase PP2A strikes back.

Therapies targeting tyrosine and serine/threonine kinases have raised enormous interest as potential cure for cancer patients in many common cancer types. However, except for the success story with BCR/ABL tyrosine kinase inhibitors in chronic myeloid leukemia (CML), critical review of results of a large number of clinical trials indicates that the clinical success with kinase inhibitors has been overall disappointing. These alarming results call for critical assessment of whether there is some fundamental flaw in the design of strategies to target phosphorylation-dependent oncogenic signaling for cancer therapy. This viewpoint debates on one potential, but thus far largely neglected, molecular explanation why inhibition of protein kinases is not sufficient for cancer cure. We note that the phosphorylation status, and thus the oncogenic potential of any given protein, is not regulated only by kinases, but rather by an intimate balance between kinases and their antagonist phosphatases. We further review the supporting functional evidence that for oncogenic transformation of human cells it is not enough to activate kinase signaling by activated kinases, if a group of counteracting tumor suppressor phosphatases is not inactivated. Based on these considerations, and a very recently emerged role of oncogenic function of a group of phosphatase inhibitor proteins as human oncoproteins, we propose that in order to efficiently inhibit phosphorylation-dependent signaling in cancer cells, and thus provide better therapeutic index, the kinase inhibitors should be combined with strategies to reactivate tumor suppressor phosphatases such as Protein Phosphatase 2A (PP2A).

Leucine Carboxyl Methyltransferase Downregulation and Protein Phosphatase Methylesterase Upregulation Contribute Toward the Inhibition of Protein Phosphatase 2A by α-Synuclein.

The pathology of Parkinson's disease (PD) is characterized by intracellular neurofibrillary tangles of phosphorylated α-synuclein (α-syn). Protein phosphatase 2A (PP2A) is responsible for α-syn dephosphorylation. Previous work has demonstrated that α-syn can regulate PP2A activity. However, the mechanisms underlying α-syn regulation of PP2A activity are not well understood. In this study, we found that α-syn overexpression induced increased α-syn phosphorylation at serine 129 (Ser129), and PP2A inhibition, in vitro and in vivo. α-syn overexpression resulted in PP2A demethylation. This demethylation was mediated via downregulated leucine carboxyl methyltransferase (LCMT-1) expression, and upregulated protein phosphatase methylesterase (PME-1) expression. Furthermore, LCMT-1 overexpression, or PME-1 inhibition, reversed α-syn-induced increases in α-syn phosphorylation and apoptosis. In addition to post-translational modifications of the catalytic subunit, regulatory subunits are involved in the regulation of PP2A activity. We found that the levels of regulatory subunits which belong to the PPP2R2 subfamily, not the PPP2R5 subfamily, were downregulated in the examined brain regions of transgenic mice. Our work identifies a novel mechanism to explain how α-syn regulates PP2A activity, and provides the optimization of PP2A methylation as a new target for PD treatment.

Non-genomic mechanisms of protein phosphatase 2A (PP2A) regulation in cancer.

Propagation of transient signals requires coordinated suppression of antagonistic phosphatase activity. Protein phosphatase 2A (PP2A) is a broad specificity serine/threonine phosphatase that functions as an antagonist of many signaling pathways associated with growth and proliferation, and endogenous inhibitory mechanisms suppress PP2A activity in response to mitogenic stimuli. These inhibitory mechanisms, including expression and activation of endogenous inhibitor proteins and phosphoregulation of PP2A subunits, are also engaged by aberrant constitutive activation of mitogenic pathways in cancer. Inhibition of PP2A activity has been shown to promote malignant transformation and endogenous inhibitory mechanisms of PP2A have been associated with malignant progression and prognosis in a wide range of cancers. Despite existence of recurrent mutations and other genetic and gene regulatory alterationsin PP2A genes, they collectively appear at relatively low frequency, and in only some cancer types. The non-genomic inhibition of PP2A activity by increased expression of endogenous PP2A inhibitor proteins greatly exceeds the frequency of genetic mutations of PP2A genes in human cancers. This feature makes PP2A an untypical tumor suppressor, and may have influenced its recognition as one of the critical human cell transformation mechanisms. We propose that non-genetic inhibition is the dominant mechanism causing loss of PP2A tumor suppressor function in cancer cells, possibly because these mechanisms do not elicit genomic instability associated with genetic loss of function of specific PP2A subunits.

Therapeutic targeting of PP2A.

Protein phosphatase 2A (PP2A) is a major serine/threonine phosphatase that regulates many cellular processes. Given the central role of PP2A in regulating diverse biological functions and its dysregulation in many diseases, including cancer, PP2A directed therapeutics have become of great interest. The main approaches leveraged thus far can be categorized as follows: 1) inhibiting endogenous inhibitors of PP2A, 2) targeted disruption of post translational modifications on PP2A subunits, or 3) direct targeting of PP2A. Additional insight into the structural, molecular, and biological framework driving the efficacy of these therapeutic strategies will provide a foundation for the refinement and development of novel and clinically tractable PP2A targeted therapies.

Downregulation of protein phosphatase 2A by apolipoprotein E: Implications for Alzheimer's disease.

The apolipoprotein E ε4 allele is the single most important genetic risk factor associated with Alzheimer's disease (AD). Tau phosphorylation and hyperphosphorylation is an underlying feature of AD and is regulated by specific kinases and phosphatases. Among phosphatases, protein phosphatase 2A (PP2A) is the principal tau dephosphorylating enzyme in the brain. Several abnormalities of PP2A have been reported in AD, including among others decreased protein levels of PP2A, decreased mRNA and protein levels of the catalytic subunit PP2AC and variable regulatory B subunits and reduced methylation of the catalytic subunit, all of which results in disruption of the PP2A phosphatase activity. In earlier studies we described a novel mechanism for ApoE as a transcription factor that binds regions of double-stranded DNA with high affinity, including the promoter regions of ~3000 different genes. The list of genes also included PPP2R5E (B56ε), a regulatory B' subunit of protein phosphatase 2A. Using a combination of A172 human glioblastoma cells, ApoE3/4 and ApoE-/- NSC and human postmortem tissue, we now demonstrate that ApoE not only binds to the PPP2R5E promoter but also triggers a significant reduction in PP2A activity by two mechanisms: 1) ApoE transcriptionally represses PPP2R5E and reduces protein expression, and 2) ApoE triggers demethylation of the catalytic subunit (PP2AC) of PP2A, resulting in the disruption of the PPP2R5E-PP2AC complex. Our results indicated a significant down-regulation of PPP2R5E gene expression and reduction in PP2A activity by ApoE4 compared with ApoE3. This may also explain an elevated Tau phosphorylation in AD human brains that featured at least one ApoE4 allele. Thus, our present work links ApoE and PPP2R5E expression to a reduction in the PP2A catalytic activity that has implications for Alzheimer's disease.

Light regulation of nitrate reductase by catalytic subunits of protein phosphatase 2A.

MAIN CONCLUSION: PP2A catalytic subunit C2 is of special importance for light/dark regulation of nitrate reductase activity. The level of unmethylated PP2A catalytic subunits decreases in darkness. Protein phosphatase 2A (PP2A) dephosphorylates and activates nitrate reductase (NR) in photosynthetically active tissue when plants are transferred from darkness to light. In the present work, investigation of Arabidopsis thaliana PP2A mutant lines revealed that one of the five PP2A catalytic subunit genes, e.g., C2, was of special importance for NR activation. Impairment of NR activation was, especially pronounced in the c2c4 double mutant. Though weaker, NR activation was also impaired in the c2 single mutant, and c1c2 and c2c5 double mutants. On the other hand, NR activation in the c4c5 double mutant was as efficient as in WT. The c4 single mutant had low PP2A activity, whereas the c2 single mutant possessed WT levels of extractable PP2A activity. PP2A activity was low in both c2c4 and c4c5. Differences in extracted PP2A activity among mutants did not strictly correlate with differences in NR activation, but underpinned that C2 has a special function in NR activation in vivo. The terminal leucine in PP2A catalytic subunits is generally methylated to a high degree, but regulation and impact of methylation/demethylation is barely studied. In WT and PP2A mutants, the level of unmethylated PP2A catalytic subunits decreased during 45 min of darkness, but did not change much when light was switched on. In leucine carboxyl methyl transferase1 (LCMT1) knockout plants, which possess mainly unmethylated PP2A, NR was still activated, although not fully as efficient as in WT.

Knockdown of microRNA-195 contributes to protein phosphatase-2A inactivation in rats with chronic brain hypoperfusion.

Reduction of protein phosphatase-2A (PP2A) activity is a common clinical feature of Alzheimer's disease and vascular dementia. In this study, we observed that chronic brain hypoperfusion induced by bilateral common carotid artery occlusion of rats led to PP2A inactivation based on the increase in tyrosine-307 phosphorylation and leucine-309 demethylation of PP2AC and the depression in PP2ABα. Knockdown of miR-195 using overexpression of its antisense molecule oligonucleotide (pre-AMO-miR-195) delivered by a lentivirus (lenti-pre-AMO-miR-195) increased tyrosine-307 phosphorylation and decreased both PP2ABα expression and leucine-309 methylation; these effects were prevented by the overexpression of miR-195 using lenti-pre-miR-195 and controlled by an increase in methylesterase (PME-1) and a decrease in leucine carboxyl methyltransferase-1. In vitro studies demonstrated that miR-195 regulated PME-1 expression by binding to the Ppme1 gene 3'-untranslated region (3'UTR) domain. Masking the miR-195 binding sites in the amyloid precursor protein (APP) and ß-site APP cleaving enzyme 1 genes prevented miR-195-induced leucine carboxyl methyltransferase-1 elevation. We concluded that the miR-195 downregulation in chronic brain hypoperfusion involved PP2A inactivity, which was mediated by the post-transcriptional regulation PME-1, APP, and ß-site APP cleaving enzyme 1 expression.

Inhibition of protein methylesterase 1 decreased cancerous phenotypes in endometrial adenocarcinoma cell lines and xenograft tumor models.

Protein methylesterase 1 (PME-1) promotes cancerous phenotypes through the demethylation and inactivation of protein phosphatase 2A. We previously demonstrated that PME-1 overexpression promotes Akt, ERK, and may promote Wnt signaling and increases tumor burden in a xenograft model of endometrial cancer. Here, we show that covalent PME-1 inhibitors decrease cell proliferation and invasive growth in vitro but have no effect in vivo at the concentrations tested; however, depletion of PME-1 with shRNA in an endometrial cancer xenograft model significantly reduced tumor growth. Thus, discovery of more potent PME-1 inhibitors may be beneficial for the treatment of endometrial cancer.

Protein phosphatase methylesterase-1 (PME-1) expression predicts a favorable clinical outcome in colorectal cancer.

Colorectal cancer (CRC) accounts for high mortality. So far, there is lack of markers capable of predicting which patients are at risk of aggressive course of the disease. Protein phosphatase-2A (PP2A) inhibitor proteins have recently gained interest as markers of more aggressive disease in certain cancers. Here, we report the role of PP2A inhibitor PME-1 in CRC. PME-1 expression was assessed from a rectal cancer patient cohort by immunohistochemistry, and correlations were performed for various clinicopathological variables and patient survival. Rectal cancer patients with higher cytoplasmic PME-1 protein expression (above median) had less recurrences (P = 0.003, n = 195) and better disease-free survival (DFS) than the patients with low cytoplasmic PME-1 protein expression (below median). Analysis of PPME-1 mRNA expression from TCGA dataset of colon and rectal adenocarcinoma (COADREAD) patient cohort confirmed high PPME1 expression as an independent protective factor predicting favorable overall survival (OS) (P = 0.005, n = 396) compared to patients with low PPME1 expression. CRC cell lines were used to study the effect of PME-1 knockdown by siRNA on cell survival. Contrary to other cancer types, PME-1 inhibition in CRC cell lines did not reduce the viability of cells or the expression of active phosphorylated AKT and ERK proteins. In conclusion, PME-1 expression predicts for a favorable outcome of CRC patients. The unexpected role of PME-1 in CRC in contrast with the oncogenic role of PP2A inhibitor proteins in other malignancies warrants further studies of cancer-specific function for each of these proteins.