The Experts below are selected from a list of 75882 Experts worldwide ranked by ideXlab platform
Isabella Russo - One of the best experts on this subject based on the ideXlab platform.
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leucine rich repeat kinase 2 controls protein kinase a activation state through phosphodiesterase 4
Journal of Neuroinflammation, 2018Co-Authors: Isabella Russo, Giulietta Di Benedetto, Alice Kaganovich, Jinhui Ding, Daniela Mercatelli, Michele Morari, Mark R Cookson, Luigi Bubacco, Elisa GreggioAbstract:Evidence indicates a cross-regulation between two kinases, leucine-rich repeat kinase 2 (LRRK2) and protein kinase A (pKa). In neurons, LRRK2 negatively regulates pKa activity in spiny projecting neurons during synaptogenesis and in response to dopamine D1 receptor activation acting as an A-anchoring kinase protein (AKAP). In microglia cells, we showed that LRRK2 kinase activity negatively regulates pKa, impacting NF-κB p50 signaling and the inflammatory response. Here, we explore the molecular mechanism underlying the functional interaction between LRRK2 and pKa in microglia. To understand which step of pKa signaling is modulated by LRRK2, we used a combination of in vitro and ex vivo systems with hyperactive or inactive LRRK2 as well as different readouts of pKa signaling. We confirmed that LRRK2 kinase activity acts as a negative regulator of pKa activation state in microglia. Specifically, we found that LRRK2 controls pKa by affecting phosphodiesterase 4 (PDE4) activity, modulating cAMP degradation, content, and its dependent signaling. Moreover, we showed that LRRK2 carrying the G2019S pathological mutation downregulates pKa activation causing a reduction of pKa-mediated NF-κB inhibitory signaling, which results, in turn, in increased inflammation in LRRK2 G2019S primary microglia upon α-synuclein pre-formed fibrils priming. Overall, our findings indicate that LRRK2 kinase activity is a key regulator of pKa signaling and suggest PDE4 as a putative LRRK2 effector in microglia. In addition, our observations suggest that LRRK2 G2019S may favor the transition of microglia toward an overactive state, which could widely contribute to the progression of the pathology in LRRK2-related PD.
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Leucine-rich repeat kinase 2 controls protein kinase A activation state through phosphodiesterase 4
'Springer Science and Business Media LLC', 2018Co-Authors: Isabella Russo, Giulietta Di Benedetto, Alice Kaganovich, Jinhui Ding, Daniela Mercatelli, Michele Morari, Mark R Cookson, Luigi Bubacco, Elisa GreggioAbstract:Abstract Background Evidence indicates a cross-regulation between two kinases, leucine-rich repeat kinase 2 (LRRK2) and protein kinase A (pKa). In neurons, LRRK2 negatively regulates pKa activity in spiny projecting neurons during synaptogenesis and in response to dopamine D1 receptor activation acting as an A-anchoring kinase protein (AKAP). In microglia cells, we showed that LRRK2 kinase activity negatively regulates pKa, impacting NF-κB p50 signaling and the inflammatory response. Here, we explore the molecular mechanism underlying the functional interaction between LRRK2 and pKa in microglia. Methods To understand which step of pKa signaling is modulated by LRRK2, we used a combination of in vitro and ex vivo systems with hyperactive or inactive LRRK2 as well as different readouts of pKa signaling. Results We confirmed that LRRK2 kinase activity acts as a negative regulator of pKa activation state in microglia. Specifically, we found that LRRK2 controls pKa by affecting phosphodiesterase 4 (PDE4) activity, modulating cAMP degradation, content, and its dependent signaling. Moreover, we showed that LRRK2 carrying the G2019S pathological mutation downregulates pKa activation causing a reduction of pKa-mediated NF-κB inhibitory signaling, which results, in turn, in increased inflammation in LRRK2 G2019S primary microglia upon α-synuclein pre-formed fibrils priming. Conclusions Overall, our findings indicate that LRRK2 kinase activity is a key regulator of pKa signaling and suggest PDE4 as a putative LRRK2 effector in microglia. In addition, our observations suggest that LRRK2 G2019S may favor the transition of microglia toward an overactive state, which could widely contribute to the progression of the pathology in LRRK2-related PD
Elisa Greggio - One of the best experts on this subject based on the ideXlab platform.
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leucine rich repeat kinase 2 controls protein kinase a activation state through phosphodiesterase 4
Journal of Neuroinflammation, 2018Co-Authors: Isabella Russo, Giulietta Di Benedetto, Alice Kaganovich, Jinhui Ding, Daniela Mercatelli, Michele Morari, Mark R Cookson, Luigi Bubacco, Elisa GreggioAbstract:Evidence indicates a cross-regulation between two kinases, leucine-rich repeat kinase 2 (LRRK2) and protein kinase A (pKa). In neurons, LRRK2 negatively regulates pKa activity in spiny projecting neurons during synaptogenesis and in response to dopamine D1 receptor activation acting as an A-anchoring kinase protein (AKAP). In microglia cells, we showed that LRRK2 kinase activity negatively regulates pKa, impacting NF-κB p50 signaling and the inflammatory response. Here, we explore the molecular mechanism underlying the functional interaction between LRRK2 and pKa in microglia. To understand which step of pKa signaling is modulated by LRRK2, we used a combination of in vitro and ex vivo systems with hyperactive or inactive LRRK2 as well as different readouts of pKa signaling. We confirmed that LRRK2 kinase activity acts as a negative regulator of pKa activation state in microglia. Specifically, we found that LRRK2 controls pKa by affecting phosphodiesterase 4 (PDE4) activity, modulating cAMP degradation, content, and its dependent signaling. Moreover, we showed that LRRK2 carrying the G2019S pathological mutation downregulates pKa activation causing a reduction of pKa-mediated NF-κB inhibitory signaling, which results, in turn, in increased inflammation in LRRK2 G2019S primary microglia upon α-synuclein pre-formed fibrils priming. Overall, our findings indicate that LRRK2 kinase activity is a key regulator of pKa signaling and suggest PDE4 as a putative LRRK2 effector in microglia. In addition, our observations suggest that LRRK2 G2019S may favor the transition of microglia toward an overactive state, which could widely contribute to the progression of the pathology in LRRK2-related PD.
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Leucine-rich repeat kinase 2 controls protein kinase A activation state through phosphodiesterase 4
'Springer Science and Business Media LLC', 2018Co-Authors: Isabella Russo, Giulietta Di Benedetto, Alice Kaganovich, Jinhui Ding, Daniela Mercatelli, Michele Morari, Mark R Cookson, Luigi Bubacco, Elisa GreggioAbstract:Abstract Background Evidence indicates a cross-regulation between two kinases, leucine-rich repeat kinase 2 (LRRK2) and protein kinase A (pKa). In neurons, LRRK2 negatively regulates pKa activity in spiny projecting neurons during synaptogenesis and in response to dopamine D1 receptor activation acting as an A-anchoring kinase protein (AKAP). In microglia cells, we showed that LRRK2 kinase activity negatively regulates pKa, impacting NF-κB p50 signaling and the inflammatory response. Here, we explore the molecular mechanism underlying the functional interaction between LRRK2 and pKa in microglia. Methods To understand which step of pKa signaling is modulated by LRRK2, we used a combination of in vitro and ex vivo systems with hyperactive or inactive LRRK2 as well as different readouts of pKa signaling. Results We confirmed that LRRK2 kinase activity acts as a negative regulator of pKa activation state in microglia. Specifically, we found that LRRK2 controls pKa by affecting phosphodiesterase 4 (PDE4) activity, modulating cAMP degradation, content, and its dependent signaling. Moreover, we showed that LRRK2 carrying the G2019S pathological mutation downregulates pKa activation causing a reduction of pKa-mediated NF-κB inhibitory signaling, which results, in turn, in increased inflammation in LRRK2 G2019S primary microglia upon α-synuclein pre-formed fibrils priming. Conclusions Overall, our findings indicate that LRRK2 kinase activity is a key regulator of pKa signaling and suggest PDE4 as a putative LRRK2 effector in microglia. In addition, our observations suggest that LRRK2 G2019S may favor the transition of microglia toward an overactive state, which could widely contribute to the progression of the pathology in LRRK2-related PD
Jean Sirois - One of the best experts on this subject based on the ideXlab platform.
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role of upstream stimulatory factor phosphorylation in the regulation of the prostaglandin g h synthase 2 promoter in granulosa cells
Journal of Biological Chemistry, 2005Co-Authors: Khampoune Sayasith, Jacques G Lussier, Jean SiroisAbstract:Abstract To investigate the role of USF phosphorylation in the regulation of the PGHS-2 promoter in granulosa cells, promoter activity assays were performed in primary cultures of bovine granulosa cells transfected with the chimeric PGHS-2 promoter/luciferase (LUC) construct –149/–2PGHS-2.LUC. Transfections were done in the absence or presence of forskolin; the protein kinase A (pKa) inhibitor H-89; or an expression vector encoding USF1, USF2, the catalytic subunit of pKa (cpKa), or a pKa inhibitor protein (PKI). Electrophoretic mobility shift assays were performed to study USF/DNA interactions using granulosa cell nuclear extracts and a 32P-labeled proximal PGHS-2 promoter fragment containing the E-box element. The results show that forskolin stimulation and cpKa overexpression caused a marked and significant increase in USF-dependent DNA binding and PGHS-2 promoter activities (p < 0.05). In contrast, both activities were decreased by H-89 treatment or PKI overexpression. Reverse transcription-PCR analyses revealed that these treatments had similar effects on endogenous PGHS-2 mRNA levels in granulosa cells. Cotransfection studies with a USF2 mutant lacking N-terminal activation domains (U2Δ1–220) repressed forskolin-, cpKa-, and USF-dependent PGHS-2 promoter activities. Electrophoretic mobility shift assays showed that U2Δ1–220 was able to compete with full-length USF proteins and to saturate the E-box element. Immunoprecipitation/Western blot analyses revealed an increase in the levels of phosphorylated USF1 and USF2 after forskolin treatment, whereas chromatin immunoprecipitation assays showed that binding of USF proteins to the endogenous PGHS-2 promoter was stimulated by forskolin. Site-directed mutagenesis of a consensus pKa phosphorylation site within USF proteins abolished their transactivating capacity. Collectively, these results characterize the role of USF phosphorylation in PGHS-2 expression and identify the phosphorylation-dependent increase in USF binding to the E-box as a putative molecular basis for the increase in PGHS-2 promoter transactivation in granulosa cells.
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role of upstream stimulatory factor phosphorylation in the regulation of the prostaglandin g h synthase 2 promoter in granulosa cells
Journal of Biological Chemistry, 2005Co-Authors: Khampoune Sayasith, Jacques G Lussier, Jean SiroisAbstract:To investigate the role of USF phosphorylation in the regulation of the PGHS-2 promoter in granulosa cells, promoter activity assays were performed in primary cultures of bovine granulosa cells transfected with the chimeric PGHS-2 promoter/luciferase (LUC) construct –149/–2PGHS-2.LUC. Transfections were done in the absence or presence of forskolin; the protein kinase A (pKa) inhibitor H-89; or an expression vector encoding USF1, USF2, the catalytic subunit of pKa (cpKa), or a pKa inhibitor protein (PKI). Electrophoretic mobility shift assays were performed to study USF/DNA interactions using granulosa cell nuclear extracts and a 32P-labeled proximal PGHS-2 promoter fragment containing the E-box element. The results show that forskolin stimulation and cpKa overexpression caused a marked and significant increase in USF-dependent DNA binding and PGHS-2 promoter activities (p
Diane M Simeone - One of the best experts on this subject based on the ideXlab platform.
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protein kinase a modulates transforming growth factor β signaling through a direct interaction with smad4 protein
Journal of Biological Chemistry, 2013Co-Authors: Huibin Yang, Lidong Wang, Michael D Uhler, Diane M SimeoneAbstract:Transforming growth factor β (TGFβ) signaling normally functions to regulate embryonic development and cellular homeostasis. It is increasingly recognized that TGFβ signaling is regulated by cross-talk with other signaling pathways. We previously reported that TGFβ activates protein kinase A (pKa) independent of cAMP through an interaction of an activated Smad3-Smad4 complex and the regulatory subunit of the pKa holoenzyme (pKa-R). Here we define the interaction domains of Smad4 and pKa-R and the functional consequences of this interaction. Using a series of Smad4 and pKa-R truncation mutants, we identified amino acids 290–300 of the Smad4 linker region as critical for the specific interaction of Smad4 and pKa-R. Co-immunoprecipitation assays showed that the B cAMP binding domain of pKa-R was sufficient for interaction with Smad4. Targeting of B domain regions conserved among all pKa-R isoforms and exposed on the molecular surface demonstrated that amino acids 281–285 and 320–329 were required for complex formation with Smad4. Interactions of these specific regions of Smad4 and pKa-R were necessary for TGFβ-mediated increases in pKa activity, CREB (cAMP-response element-binding protein) phosphorylation, induction of p21, and growth inhibition. Moreover, this Smad4-pKa interaction was required for TGFβ-induced epithelial mesenchymal transition, invasion of pancreatic tumor cells, and regulation of tumor growth in vivo.
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a transforming growth factor β induced smad3 smad4 complex directly activates protein kinase a
Molecular and Cellular Biology, 2004Co-Authors: Lizhi Zhang, Michael D Uhler, Chao Jun Duan, Charles E Binkley, Craig D Logsdon, Diane M SimeoneAbstract:Transforming growth factor β (TGFβ) interacts with cell surface receptors to initiate a signaling cascade critical in regulating growth, differentiation, and development of many cell types. TGFβ signaling involves activation of Smad proteins which directly regulate target gene expression. Here we show that Smad proteins also regulate gene expression by using a previously unrecognized pathway involving direct interaction with protein kinase A (pKa). pKa has numerous effects on growth, differentiation, and apoptosis, and activation of pKa is generally initiated by increased cellular cyclic AMP (cAMP). However, we found that TGFβ activates pKa independent of increased cAMP, and our observations support the conclusion that there is formation of a complex between Smad proteins and the regulatory subunit of pKa, with release of the catalytic subunit from the pKa holoenzyme. We also found that the activation of pKa was required for TGFβ activation of CREB, induction of p21Cip1, and inhibition of cell growth. Taken together, these data indicate an important and previously unrecognized interaction between the TGFβ and pKa signaling pathways.
Karan Garg - One of the best experts on this subject based on the ideXlab platform.
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statin use and other factors associated with mortality after major lower extremity amputation
Journal of Vascular Surgery, 2017Co-Authors: Charles Decarlo, John P Phair, Saadat Shariff, Evan C Lipsitz, Larry Scher, Karan GargAbstract:Abstract Objective Above-knee amputations (AKAs) and below-knee amputations (BKAs) are associated with high postoperative mortality rates. In this study, we examined factors associated with 30-day, 90-day, and 1-year mortality in patients who underwent a major lower extremity amputation. Methods We queried a prospectively collected institutional database for all patients who underwent AKA or BKA with primary or secondary closure, during a 5-year period, between November 2009 and November 2014. Predictors of 30- and 90-day mortality were determined by multivariable logistic regression, and risk indexes for 1-year mortality were determined with Cox proportional hazards model. Results We identified 811 patients who underwent AKA (n = 325) or BKA (n = 486). The 30-day mortality was 8.4% (AKA, 13.5%; BKA, 4.9%; P P 70 years (HR, 1.39; 95% CI, 1.02-1.88), gangrene (HR, 1.44; 95% CI, 1.07-1.94), ESRD on HD (HR, 1.96; 95% CI, 1.42-2.70), chronic obstructive pulmonary disease (HR, 2.54; 95% CI, 1.52-4.25), Caucasian race (HR, 1.62; 95% CI, 1.18-2.22), history of open lower extremity revascularization (HR, 0.71; 95% CI, 0.51-1.00) and undergoing bilateral amputations (HR, 2.10; 95% CI, 1.06-4.15). In the year after amputation, medium-intensity statin (HR, 0.64; 95% CI, 0.47-0.87) and high-intensity statin (HR, 0.56; 95% CI, 0.33-0.95) conferred a mortality benefit. Low-intensity statins did not confer protection from mortality. At 1 year after amputation, only 44.7% of patients were receiving appropriate statin therapy. Conclusions AKA and BKA have historically been associated with high mortality rates. Medium-intensity and high-intensity statin therapies were associated with a mortality benefit at 1 year. We have identified initiation of statin therapy in this high-risk population as a gap in patient care.
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risk factors for unplanned readmission and stump complications after major lower extremity amputation
Journal of Vascular Surgery, 2016Co-Authors: John P Phair, Charles Decarlo, Larry A Scher, Issam Koleilat, Saadat Shariff, Evan C Lipsitz, Karan GargAbstract:Abstract Objective The unplanned 30-day readmission rate is a marker of quality of patient care across many disciplines. Data regarding risk factors for unplanned readmission after major lower extremity amputation (LEA) are limited. We evaluated predictors of readmission at our institution after major LEA. Methods We conducted a retrospective review of all patients undergoing above-knee amputation (AKA) or below-knee amputation (BKA) between November 2009 and November 2014. Patient demographic variables were collected. Predictors of unplanned 30-day readmission and stump complications were determined by multivariable logistic regression. Results A total of 811 patients were identified (AKA, 325; BKA, 486). Of these, 739 patients were included in the final analysis after excluding 30-day decedents without readmission. The overall 30-day readmission rate was 28.8% (AKA 27.9%; BKA 29.4%; P = .730). Stump complications accounted for 28.6% of readmissions (16.5% of AKA; 35.8% of BKA; P = .004). Other common diagnoses included nonsurgical site infection (33.8%), exacerbation of congestive heart failure (7.0%), and diabetes-related complications (6.1%). Surgical intervention was performed on 61% of stump complications (35.9% of AKA readmitted with stump complications; 68.7% of BKA readmitted with stump complications). BKA stump complications were converted to AKAs in 34.1% of cases (3.2% of the total BKA). None of the AKA stump complications required a higher level of amputation (ie, hip disarticulation). Independent predictors of all 30-day readmission included coronary artery disease and end-stage renal disease. American Society of Anesthesiologists class 3 as compared with class 4 was protective. Independent predictors of 30-day readmission for stump complications included rest pain and BKA. Patients who underwent BKA, rest pain as an indication for amputation, and having an occluded bypass graft were predictors of having a stump complication requiring surgery. Conclusions The 30-day readmission rate after major LEA is high, with wound infections accounting for a significant proportion of these readmissions. There was no difference in readmission rates based on level of amputation. Those undergoing BKA were more likely to present with stump complications requiring a surgical intervention, and often a higher level of amputation. Identification of high-risk patients may play a role in reducing postoperative readmissions and stump complications.