Nuclear Phosphatase Research Articles

AMPK Suppresses Vascular Inflammation In Vivo by Inhibiting Signal Transducer and Activator of Transcription-1.

Activation of AMPK suppresses inflammation, but the underlying mechanisms remain poorly understood. This study was designed to characterize the molecular mechanisms by which AMPK suppresses vascular inflammation. In cultured human aortic smooth muscle cells, pharmacologic or genetic activation of AMPK inhibited the signal transducer and activator of transcription-1 (STAT1), while inhibition of AMPK had opposite effects. Deletion of AMPKα1 or AMPKα2 resulted in activation of STAT1 and in increases in proinflammatory mediators, both of which were attenuated by administration of STAT1 small interfering RNA or fludarabine, a selective STAT1 inhibitor. Moreover, AMPK activation attenuated the proinflammatory actions induced by STAT1 activators such as interferon-γ and angiotensin II (AngII). Mechanistically, we found that AMPK activation increased, whereas AMPK inhibition decreased, the levels of mitogen-activated protein kinase phosphatase-1 (MKP-1), an inducible nuclear phosphatase, by regulating proteasome-dependent degradation of MKP-1. Gene silencing of MKP-1 increased STAT1 phosphorylation and prevented 5-aminoimidazole-4-carboxyamide ribonucleoside–reduced STAT1 phosphorylation. Finally, we found that infusion of AngII caused a more severe inflammatory response in AMPKα2 knockout mouse aortas, all of which were suppressed by chronic administration of fludarabine. We conclude that AMPK activation suppresses STAT1 signaling and inhibits vascular inflammation through the upregulation of MKP-1.

Inducing malignant transformation of endometriosis in rats by long-term sustaining hyperestrogenemia and type II diabetes.

This study aimed to induce malignant transformation of endometriosis in Sprague–Dawley rats by hyperestrogenemia and type II diabetes and evaluate its similarity with human disease in biological features. Rats with surgically induced endometriosis were randomized into two groups: those treated with estradiol (5 mg/kg three times/week after surgery), streptozotocin (25 mg/kg, 1 month after surgery), and high carbohydrate-and-fat feed (Es group); and those treated with placebo saline and standard feed (control group). All rats were randomly killed 2, 4, or 8 months after surgery. The endometriosis lesions and the corresponding eutopic endometria were subjected to morphological evaluation, TUNEL, and immunohistochemical analysis for the expressions of proliferating cell nuclear antigen, phosphatase and tensin homolog, phosphorylated protein kinase B, and phosphorylated mammalian target of rapamycin proteins. In the Es group, three cases (6.0%) of endometriosis showed atypical hyperplasia accompanied by simple hyperplastic eutopic endometria, and two cases (4.0%) of endometriosis showed endometrioid carcinoma accompanied by atypical hyperplastic eutopic endometria. In the Es group, the activity of organelles and the expressions of proliferating cell nuclear antigen, phosphorylated protein kinase B, and phosphorylated mammalian target of rapamycin increased, and the level of phosphatase and tensin homolog and TUNEL positivity decreased progressively in the order of endometriosis, atypical endometriosis, and malignant endometriosis. The same tendency was found in the corresponding eutopic endometria. The induced malignant endometriosis showed similarities with human disease in the pathological process and histomorphological and molecular biological features. The method is feasible. The malignant transformations of endometriosis and eutopic endometria may have correlations and similarities, but the former may suffer a higher risk of canceration.

Abstract A05: The control of nuclear ERK activity by the DUSP phosphatases in colorectal cancer

Abstract The RAS/RAF/MEK/ERK pathway is suspected to be strongly implicated in malignant transformation of colorectal epithelial cells since KRAS and BRAF genes are mutated in up to 60% of human colorectal cancers (CRCs). Despite intensive efforts made in the understanding of KRAS and BRAF functions in colorectal carcinogenesis, little is known about the oncogenic regulation of ERK activity in CRC cells, especially by the ERK-specific nuclear DUSP phosphatases. We have recently demonstrated in CRC cells that ERK1/2 activity is confined to the cytoplasm because of a pervanadate-dependent phosphatase activity. Accordingly, DUSP4 mRNA was found to be highly expressed, in a MEK-dependent manner, in CRC cells. Thus, DUSP4 and other nuclear DUSPs may function as part of a negative feedback mechanism in the control of the duration and magnitude of nuclear ERK activation during colorectal carcinogenesis. We thus speculate that the modification of nuclear ERK activity might strongly impact on the oncogenic properties of CRC. Methods: Expression of DUSPs was analyzed by PCR in CRC cells and qPCR in human colorectal tumors from different stages. To elucidate the role of nuclear DUSPs, RNA interference was used to specifically silence their expression in human HT29 CRC cell line, a cell line exhibiting BRafV600E mutation. Inducible expression of a DUSP4 inactivated mutant (DUSP4C284S) was done in a model of IEC-6 cells that express an inducible form of BRaf oncogene (BRafV600E:ER). Results: 1- DUSP2, DUSP4 and DUSP5 mRNAs were found to be highly expressed, in a MEK-dependent manner, in CRC cells. 2- DUSP4 and DUSP5 mRNA levels were markedly increased in colorectal tumors in comparison to healthy matched adjacent tissues. 3- Silencing of DUSP 2, 4 or 5 in HT29 cells increased ERK phosphorylation levels and markedly reduced colony formation in soft agar. 4- Furthermore, expression of DUSP4C284S in BRafV600E-expressing IEC-6 cells increased ERK phosphorylation into the nucleus and markedly reduced colony formation in soft agar. Conclusion: We are currently searching by which mechanisms sustained nuclear ERK activity reduces colorectal tumor cell growth. Therefore, targeting nuclear DUSP activity could become a new strategy to treat CRC. Citation Format: Sebastien Cagnol, Nathalie Rivard. The control of nuclear ERK activity by the DUSP phosphatases in colorectal cancer. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr A05. doi: 10.1158/1557-3125.RASONC14-A05

The catalytically inactive tyrosine phosphatase HD-PTP/PTPN23 is a novel regulator of SMN complex localization.

The survival motor neuron (SMN) complex fulfils essential functions in the assembly of snRNPs, which are key components in the splicing of pre-mRNAs. Little is known about the regulation of SMN complex activity by posttranslational modification despite its complicated phosphorylation pattern. Several phosphatases had been implicated in the regulation of SMN, including the nuclear phosphatases PPM1G and PP1γ. Here we systematically screened all human phosphatase gene products for a regulatory role in the SMN complex. We used the accumulation of SMN in Cajal bodies of intact proliferating cells, which actively assemble snRNPs, as a readout for unperturbed SMN complex function. Knockdown of 29 protein phosphatases interfered with SMN accumulation in Cajal bodies, suggesting impaired SMN complex function, among those the catalytically inactive, non-receptor-type tyrosine phosphatase PTPN23/HD-PTP. Knockdown of PTPN23 also led to changes in the phosphorylation pattern of SMN without affecting the assembly of the SMN complex. We further show interaction between SMN and PTPN23 and document that PTPN23, like SMN, shuttles between nucleus and cytoplasm. Our data provide the first comprehensive screen for SMN complex regulators and establish a novel regulatory function of PTPN23 in maintaining a highly phosphorylated state of SMN, which is important for its proper function in snRNP assembly.

C-terminal Domain (CTD) Small Phosphatase-like 2 Modulates the Canonical Bone Morphogenetic Protein (BMP) Signaling and Mesenchymal Differentiation via Smad Dephosphorylation

The bone morphogenetic protein (BMP) signaling pathway regulates a wide range of cellular responses in metazoans. A key step in the canonical BMP signaling is the phosphorylation and activation of transcription factors Smad1, Smad5, and Smad8 (collectively Smad1/5/8) by the type I BMP receptors. We previously identified PPM1A as a phosphatase toward dephosphorylation of all receptor-regulated Smads (R-Smads), including Smad1/5/8. Here we report another nuclear phosphatase named SCP4/CTDSPL2, belonging to the FCP/SCP family, as a novel Smad phosphatase in the nucleus. SCP4 physically interacts with and specifically dephosphorylates Smad1/5/8, and as a result attenuates BMP-induced transcriptional responses. Knockdown of SCP4 in multipotent mesenchymal C2C12 cells leads to increased expression of BMP target genes and consequently promotes BMP-induced osteogenic differentiation. Collectively, our results demonstrate that SCP4, as a Smad phosphatase, plays a critical role in BMP-induced signaling and cellular functions.

Abstract 215: The Redox Activation of DUSP4 by N-Acetyl Cysteine Protects Endothelial Cells and the Myocardium against Oxidative Damage

Background: Redox imbalance is the primary cause for endothelial dysfunction (ED), obstructed blood flow, and subsequent heart attack and failure. Under oxidant stress, many critical proteins regulating endothelial function undergo oxidative modifications that lead to ED. Cellular levels of glutathione (GSH), the primary reducing source, can significantly regulate cell function via reversible protein thiol modification. N-Acetyl cysteine (NAC), a precursor for GSH biosynthesis, is beneficial for many vascular diseases; however, the detailed mechanism of these benefits is still not clear. Methods: We employed EPR spin-trapping, HPLC, fluorescent microscopy, immunoblotting, and qPCR of both in vitro and ex vivo experiments using either cultured cells or the Langendorff heart preparation. Results: NAC treatment increases NO generation from endothelial cells, as well as the enzyme and cofactor responsible for its production, ie eNOS and BH4. Interestingly, NAC treatment also increased the expression of DUSP4, an inducible nuclear dual-specificity phosphatase implicated in cardiovascular function. We hereby establish that DUSP4 redox modulates two important kinases (p38 and ERK1/2) of MAPK signaling pathways and provides protection against Cd2+-induced ROS damage as well as hypoxia-reoxygenation insult to endothelial cells. Furthermore, a four week oral NAC pre-treatment promotes DUSP4 both protein and mRNA expression in the rat myocardium and renders the heart less susceptible to ex vivo ischemia-reperfusion injury. Protein expression profiles in the myocardium closely mimic those observed with cultured endothelial cells. The infarct size of NAC-treated hearts is significantly reduced. The myocardial rate pressure product is much improved above vehicle treated rats. Conclusion: NAC serves as a direct antioxidant and as a regulator of transcription and translation of DUSP4, modulating the activity of its downstream effectors: ERK1/2 and p38, and thus protects against ROS-induced damage both in vitro and ex vivo. As such, NAC-derived drugs can provide a novel therapy to oxidant-induced diseases via the specific up-regulation of protective proteins such as DUSP4 and eNOS.

Application of 31P NMR spectroscopy in determining phosphatase activities and P composition in soil aggregates influenced by tillage and residue management practices

Soil phosphorus (P) composition and phosphatase activities in aggregates are essential for agricultural productivity and remain poorly understood. A field experiment was conducted from 2007 to study the effect of tillage systems (conventional tillage, T and no tillage, NT) and crop residue management (0, 50% and 100% crop residue incorporation/coverage) on P composition determined by 31P nuclear magnetic resonance (NMR) and phosphatase activities in soil aggregates (>2mm, 0.25–2mm and 0.053–0.25mm). The results showed that crop residue input influenced the concentrations of soil phosphate monoesters and diesters, alkaline phosphomonoesterase (AlP), acid phosphomonoesterase (AcP), phosphodiesterase (PD) activities, and soil aggregate stability significantly, and the addition of crop residue was significantly more effective than tillage. The NT had significantly higher soil phosphatase activities than tillage treatment but not more soil P content. The 0.25–2mm aggregates showed higher total P, organic P, concentrations of monoesters and diesters, and AlP activity. The structure equation model showed that soil aggregate stability could increase concentrations of monoesters and diesters indirectly by its direct effects on soil phosphatases. Our results suggest that NT and crop residue input could increase the P store and sustainable supply in soil aggregates and that the 0.25–2mm size aggregates may play an important role in soil organic P maintenance and transformation.

T-cell protein tyrosine phosphatase (TCPTP) regulates phosphorylation of Txk, a tyrosine kinase of the Tec family

Phosphorylation and dephosphorylation of intracellular enzymes, such as the T-cell-specific Tec family protein kinase Txk, are critically involved in the regulation of a number of cellular functions. Although PTPs located in the nucleus are limited, TCPTP is known to be one such nuclear phosphatase, due to its nuclear localization signal sequences. Here, we investigate the role of TCPTP in the regulation of Txk phosphorylation/dephosphorylation in Txk-transfected Cos7 and Jurkat cells. Nuclear-type TCPTP (TC45) was present in both the nuclear and cytoplasmic compartments of Cos7 cells transfected with TC45. Cytoplasmic-type TCPTP (TC48) was localized in the cytoplasmic compartment of Cos7 cells transfected with TC48. We observed that expression of TC45 dephosphorylated Txk in the nuclei of Cos7 cells. TC48 expression did not dephosphorylate Txk; rather, it enhanced and sustained Txk phosphorylation in the nuclei of Cos7 cells. Phosphorylation of Txk increased 60 minutes after lectin stimulation in Jurkat cells transfected with TCPTP-specific small interfering RNA (siRNA), which efficiently knocked down endogenous TCPTP. TCPTP may play a crucial role in the regulation of Txk phosphorylation status in T-cells after stimulation. Rec./11/5/2014, Acc.11/19/2014, pp240-246

Tillage effects on phosphorus composition and phosphatase activities in soil aggregates

Phosphorus (P) and phosphatase activities in soil aggregates affected by tillage under cold monsoon climate remain poorly understood. Based on the hypothesis that the distribution of P composition and phosphatase activities in soil aggregates should be affected by different tillage practices, a field experiment was conducted to study the effects of moldboard plow (MP), ridge tillage (RT), and no-tillage (NT) on the distribution of soil P composition determined by 31P nuclear magnetic resonance (NMR) and phosphatase activities in different size fractions of soil aggregates (>2, 1–2, 0.25–1, and <0.25mm) at the 0 to 20cm depth in northeastern China. NT treatment had significantly higher organic P proportion in total P and larger proportions of monoesters and diesters in extracted total P than the MP treatment, whereas the MP treatment showed higher concentrations of total P, organic P, plant available P, NaOH-EDTA extracted total P, orthophosphate and monoesters. Soil alkaline phosphatase (AlP) and phosphodiesterase (PD) activities under NT were significantly higher than those under MP, and the responses of AlP in 0.25–1mm size fraction and PD in <0.25mm size fraction were more sensitive to tillage treatments. Overall, although NT facilitated more P stored in the organic P form and increased phosphatase activities, soil with NT had lower total and plant available P compared to traditional MP treatment and therefore, MP may be the right practice to conserve soil P under cold monsoon climate.

Oxidative stress impairs multiple regulatory events to drive persistent cytokine-stimulated STAT3 phosphorylation

Although cytokine-driven STAT3 phosphorylation and activation are often transient, persistent activation of STAT3 is a hallmark of a range of pathologies and underpins altered transcriptional responses. As triggers in disease frequently include combined increases in inflammatory cytokine and reactive oxygen species levels, we report here how oxidative stress impacts on cytokine-driven STAT3 signal transduction events. In the model system of murine embryonic fibroblasts (MEFs), combined treatment with the interleukin-6 family cytokine Leukemia Inhibitory Factor (LIF) and hydrogen peroxide (H2O2) drove persistent STAT3 phosphorylation whereas STAT3 phosphorylation increased only transiently in response to LIF alone and was not increased by H2O2 alone. Surprisingly, increases in transcript levels of the direct STAT3 gene target SOCS3 were delayed during the combined LIF+H2O2 treatment, leading us to probe the impact of oxidative stress on STAT3 regulatory events. Indeed, LIF+H2O2 prolonged JAK activation, delayed STAT3 nuclear localisation, and caused relocalisation of nuclear STAT3 phosphatase TC-PTP (TC45) to the cytoplasm. In exploring the nuclear import/export pathways, we observed disruption of nuclear/cytoplasmic distributions of Ran and importin-α3 in cells exposed to H2O2 and the resultant reduced nuclear trafficking of classical importin-α/β-dependent protein cargoes. CRM1-mediated nuclear export persisted despite the oxidative stress insult, with sustained STAT3 Y705 phosphorylation enhancing STAT3 nuclear residency. Our studies thus reveal for the first time the striking impact of oxidative stress to sustain STAT3 phosphorylation and nuclear retention following disruption of multiple regulatory events, with significant implications for STAT3 function.

Nuclear trafficking of Pten after brain injury leads to neuron survival not death

There is controversy whether accumulation of the tumor suppressor PTEN protein in the cell nucleus under stress conditions such as trauma and stroke causes cell death. A number of in vitro studies have reported enhanced apoptosis in neurons possessing nuclear PTEN, with the interpretation that its nuclear phosphatase activity leads to reduction of the survival protein phospho-Akt. However, there have been no in vivo studies to show that nuclear PTEN in neurons under stress is detrimental. Using a mouse model of injury, we demonstrate here that brain trauma altered the nucleo-cytoplasmic distribution of Pten, resulting in increased nuclear Pten but only in surviving neurons near the lesion. This event was driven by Ndfip1, an adaptor and activator of protein ubiquitination by Nedd4 E3 ligases. Neurons next to the lesion with nuclear PTEN were invariably negative for TUNEL, a marker for cell death. These neurons also showed increased Ndfip1 which we previously showed to be associated with neuron survival. Biochemical assays revealed that overall levels of Pten in the affected cortex were unchanged after trauma, suggesting that Pten abundance globally had not increased but rather Pten subcellular location in affected neurons had changed. Following experimental injury, the number of neurons with nuclear Pten was reduced in heterozygous mice (Ndfip1+/−) although lesion volumes were increased. We conclude that nuclear trafficking of Pten following injury leads to neuron survival not death.

Mitogen‐Activated Protein Kinase Phosphatase 1 as an Inflammatory Factor and Drug Target

Mitogen-activated protein kinases (MAPKs) are signaling proteins that are activated through phosphorylation, and they regulate many physiological and pathophysiological processes in cells. Mitogen-activated protein kinase phosphatase 1 (MKP-1) is an inducible nuclear phosphatase that dephosphorylates MAPKs, and thus, it is a negative feedback regulator of MAPK activity. MKP-1 has been found as a key endogenous suppressor of innate immune responses, as well as a regulator of the onset and course of adaptive immune responses. Altered MKP-1 signaling is implicated in chronic inflammatory diseases in man. Interestingly, MKP-1 expression and protein function have been found to be regulated by certain anti-inflammatory drugs, namely by glucocorticoids, antirheumatic gold compounds and PDE4 inhibitors, and MKP-1 has been shown to mediate many of their anti-inflammatory effects. In this Mini Review, we summarize the effect of MKP-1 in the regulation of innate and adaptive immune responses and its role as a potential anti-inflammatory drug target and review recent findings concerning the role of MKP-1 in certain anti-inflammatory drug effects.

Ovarian cancers arising from endometriosis: A microenvironmental biomarker study including ER, HNF1ß, p53, PTEN, BAF250a, and COX-2

BackgroundThe microenvironmental biomarkers of different subtypes of ovarian cancers arising from endometriosis have not been studied in Taiwan. Their expression can help in understanding the carcinogenic mechanism. MethodsOur study used immunohistochemistry to compare the expression of estrogen receptor (ER), hepatocyte nuclear factor-1 beta (HNF1ß), p53, phosphatase and tensin homolog (PTEN), BAF250a, and cyclooxygenase-2 (COX-2) among 79 cases of endometriosis-associated ovarian cancers, including 40 (50%) clear cell carcinomas (CCCs), 33 (41%) endometrioid (EM) adenocarcinomas, four (5%) serous carcinomas, one adenosquamous carcinoma, and one adenosarcoma. ResultsPositive stainings for ER, HNF1ß, p53, and COX-2 were identified in 34 (43%), 30 (38%), 10 (13%), and 44 (56%) cases. Loss of PTEN and BAF250a were noted in 29 (37%) and 37 (47%) cases. The expression of ER was reversely correlated with that of HNF1ß (rho = −0.417, p < 0.001) and correlated with p53 (rho = 0.284, p = 0.011). ER positivity was commonly identified in EM adenocarcinomas (91%), and rarely in CCCs (8%) and serous carcinoma (0%; p < 0.001). By contrast, HNF1ß expression was frequently noted in CCCs (65%) and serous carcinomas (50%), but less in EM adenocarcinoma (6%; p < 0.001). All staining results were similar between atypical endometriosis glandular epithelium and contiguous malignant parts. Only nine cases showed 10 minor differences (10/474, 2%) in ER, HNF1ß, and BAF250a. For the staining patterns of p53, COX-2, and PTEN, there was no difference between the invasive and precursor parts. ConclusionOur results supported the suggestion that estrogen-dependent ovarian cancer arising from endometriosis is substantially more associated with EM adenocarcinoma than CCCs. The positive HNF1ß staining was a frequent finding in CCCs, but not in EM adenocarcinoma. The similar staining patterns of atypical endometriosis glandular cells with the invasive parts confirmed their precursor status.

Nuclear phosphatase PPM1G in cellular survival and neural development

PPM1G is a nuclear localized serine/threonine phosphatase implicated to be a regulator of chromatin remodeling, mRNA splicing, and DNA damage. However, its in vivo function is unknown. Here we show that ppm1g expression is highly enriched in the central nervous system during mouse and zebrafish development. ppm1g(-/-) mice were embryonic lethal with incomplete penetrance after E12.5. Rostral defects, including neural tube and craniofacial defects were observed in ppm1g(-/-) embryos associated with increased cell death in the neural epithelium. In zebrafish, loss of ppm1g also led to neural defects with aberrant neural marker gene expression. Primary fibroblasts from ppm1g(-/-) embryos failed to grow without immortalization while immortalized ppm1g(-/-) fibroblasts had increased cell death upon oxidative and genotoxic stress when compared to wild type fibroblasts. Our in vivo and in vitro studies revealed a critical role for PPM1G in normal development and cell survival.

Clinically Relevant Dimer Interface Mutants of STAT1 Transcription Factor Exhibit Differential Gene Expression

A transition from a parallel to an antiparallel dimer configuration of the transcription factor signal transducer and activator of transcription 1 (STAT1) is required for interferon (IFN)-mediated signal transduction. However, the precise molecular mechanisms linking conformational changes to target gene activation by STAT1 are still largely unknown. In the present study, we have characterized, in more detail than before, two disease-associated point mutants with amino acid substitutions at both sites of the dimer interface (F172W and T385A). First, we confirmed that IFNγ-stimulation of transfected cells led to enhanced tyrosine phosphorylation of mutant STAT1 as compared to the wild-type protein, which consequently resulted in its prolonged nuclear accumulation. Using an in vitro dephosphorylation assay, we demonstrated that, in contrast to wild-type STAT1 and similar to the F172W mutant, also T385A resisted enzymatic inactivation by the nuclear phosphatase Tc45. Transcriptional activation of IFNγ-driven endogenous target genes differed between wild-type and mutant STAT1. While expression of genes containing a single classical gamma-activated site (GAS), such as irf1, gpb1, and mig1, was virtually unaffected by the presence of either of two amino acid exchanges, induction of the cxcl10 and mcp1 gene was significantly enhanced. The latter two genes both contain an additional TTC/GAA binding motif separated by 10 bp from the palindromic GAS sequence. The transcriptional superiority of the mutants on these genes was reflected by their increased binding affinity to DNA fragments containing the identified “one-and-a-half-GAS” motif. In summary, our data demonstrate that two clinically relevant interface mutants of STAT1 exhibit gene-specific effects and point to the rather complex role of the assumed conformational shift between two different dimer configurations for efficient transcriptional regulation.

Signal-dependent repression of DUSP5 by class I HDACs controls nuclear ERK activity and cardiomyocyte hypertrophy

Cardiac hypertrophy is a strong predictor of morbidity and mortality in patients with heart failure. Small molecule histone deacetylase (HDAC) inhibitors have been shown to suppress cardiac hypertrophy through mechanisms that remain poorly understood. We report that class I HDACs function as signal-dependent repressors of cardiac hypertrophy via inhibition of the gene encoding dual-specificity phosphatase 5 (DUSP5) DUSP5, a nuclear phosphatase that negatively regulates prohypertrophic signaling by ERK1/2. Inhibition of DUSP5 by class I HDACs requires activity of the ERK kinase, mitogen-activated protein kinase kinase (MEK), revealing a self-reinforcing mechanism for promotion of cardiac ERK signaling. In cardiac myocytes treated with highly selective class I HDAC inhibitors, nuclear ERK1/2 signaling is suppressed in a manner that is absolutely dependent on DUSP5. In contrast, cytosolic ERK1/2 activation is maintained under these same conditions. Ectopic expression of DUSP5 in cardiomyocytes results in potent inhibition of agonist-dependent hypertrophy through a mechanism involving suppression of the gene program for hypertrophic growth. These findings define unique roles for class I HDACs and DUSP5 as integral components of a regulatory signaling circuit that controls cardiac hypertrophy.

Final steps in the feedback regulation of human glucocorticoid receptor gene and role of nuclear protein phosphatase 2A

Abstract The feedback regulation of the human glucocorticoid receptor gene by glucocorticoid receptor-hormone complexes involves the recruitment of the nuclear phosphatase, PP2A. The PP2A holoenzyme consists of a structural subunit A, a

Identification of a novel cell-penetrating peptide from human phosphatidate phosphatase LPIN3.

Biomolecules such as proteins, DNA, and RNA are macromolecules and can not cross the cell membrane. However, cell-penetrating peptide (CPP) has been shown to deliver therapeutic biomolecules successfully into cells. The various and widely used CPPs including TAT, VP22, and Antp are mostly non-human originated CPPs, and are limited by their potential toxicity and immunogenicity. We report here on a newly identified novel cell-penetrating sequence (LPIN; RRKRRRRRK) from the nuclear localization sequence (NLS) of human nuclear phosphatase, LPIN3. LPIN-EGFP recombinant protein was concentration- and time-dependently delivered into cells and localized to the nucleus as well as the cytoplasm. It penetrated the cell membrane by lipid raft-mediated endocytosis by binding to heparan sulfate proteoglycan. LPIN-EGFP was successfully delivered into primary mouse splenocytes in vitro and it could be delivered into various tissues including liver, kidney, and intestine in mice after intra-peritoneal injection. This research suggests that LPIN-CPP could be used in a drug delivery system to deliver therapeutic biomolecules including peptides, proteins, DNA, and RNA and without the limitations of non-human originated CPPs such as TAT-CPP.

A Novel Nuclear Protein Phosphatase 2C Negatively Regulated by ABL1 is Involved in Abiotic Stress and Panicle Development in Rice

Type 2C protein phosphatase plays an important role in the signal transduction of stress response in plants. In this paper, we identified a novel stress-induced type 2C protein phosphatase gene OsSIPP2C1 from rice. OsSIPP2C1 contains a complete open reading frame of 1,074bp, encoding a protein with 357 amino acids. OsSIPP2C1 expression was up-regulated by high salt, PEG6000 and exogenous ABA, and enhanced in the abl1 mutant under normal, salt, or drought condition. Interestingly, OsSIPP2C1 expression was increased during the early panicle development. Subcellular localization assay using rice protoplast cells indicated that OsSIPP2C1 was predominantly located in the nucleus. Together, it is suggested that a nuclear PP2C protein OsSIPP2C1 negatively regulated by ABL1 is involved in abiotic stress and panicle development in rice.

Regulation of tristetraprolin expression by mitogen-activated protein kinase phosphatase-1

Tristetraprolin (TTP) is an acute phase protein, and its expression is rapidly up-regulated by inflammatory signals, such as lipopolysaccharide (LPS) and cytokines. TTP regulates gene expression by governing the mRNA stability of its target genes, which include cytokines and growth factors. MAP kinase phosphatase-1 (MKP-1) is a nuclear phosphatase that inhibits p38 mitogen-activated protein kinase (MAPK) signaling. This study investigated the role of MKP-1 in TTP expression in A549 human lung epithelial cells, THP-1 human macrophages, J774 mouse macrophages, and primary mouse macrophages. TTP and MKP-1 expression was increased by cytokines or LPS. Silencing of MKP-1 by siRNA enhanced TTP expression in response to LPS, and LPS-induced TTP expression was increased in macrophages from MKP-1 (-/-) mice as compared with that in macrophages from wild-type animals. The inhibition of p38 MAPK by SB202190 reduced TTP expression. In conclusion, MKP-1 suppressed TTP expression by inhibiting p38 MAPK pathway.