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University of Rochester

1. Madukwe, Jerry Chukwuemeka. Insights into G Protein βγ Regulation of Phospholipase Cε.

Degree: PhD, 2020, University of Rochester

PLCϵ is involved in the regulation of multiple cellular signaling processes and it has been implicated in the development of heart failure. Due to its central importance, an intimate understanding of its regulation is critical for developing potential pharmacological therapies. To identify domains of Phospholipase Cϵ (PLCϵ) that interact with G protein βγ (Gβγ), we designed various point mutations and truncations of wild-type PLCϵ and tested them for Gβγ activation in transfected COS-7 cells. In contrast to activation by Ras, deletion of an individual domain in PLCϵ is not sufficient to completely block Gβγ mediated activation. Simultaneous deletion of the carboxy-terminal RA2 domain and the amino-terminal (CDC25 and cysteine-rich domains) results in complete abrogation of PLCϵ activation by Gβγ, while activation by Rho is retained. In vitro reconstitution experiments revealed that purified Gβγ directly interacts with a purified fragment of PLCϵ (PLCϵ-PH-RA2) to increase membrane phosphatidylinositol 4,5 bisphosphate (PIP2) hydrolysis. Deletion of the RA2 domain decreased Gβγ binding and eliminated Gβγ-stimulation of PIP2 hydrolysis. Our data show that two independent domains of PLCϵ are involved in its regulation by Gβγ and demonstrate direct interactions between Gβγ and PLCϵ. Furthermore, to clearly understand the mechanism of Gβγ regulation of Golgi compartmentalized PLCϵ (Golgi-PLCϵ) by plasma membrane (PM) endothelin receptor (ET-1AR) activation, which is required for cardiomyocyte hypertrophy in a neonatal rat ventricular myocyte (NRVM) model of cardiac disease, we overexpressed differentially translocating Gγ in NRVMs using adenovirus. We tested if G protein γ (Gγ)-mediated differential translocation of Gβγ differentially modulates Gβγ dependent Golgi-PLCϵ-mediated phosphatidylinositol 4-phosphate (PI4P) hydrolysis upon stimulation of the ET-1AR. Our data show that Gγ differences in translocation characteristics do not confer any differential effect in Golgi-PLCϵ-mediated PI4P hydrolysis upon ET-1AR activation. Finally, we explored differential modulation by Gγ of cardiac cell hypertrophy and protein kinase D (PKD); a downstream effector of Gβγ dependent Golgi-PLCϵ signaling that regulates cardiac hypertrophy. Our data show that Gγ-mediated differential translocation of Gβγ confers no differential effect on PKD activation and cellular hypertrophy. Taken together, results of our studies provide the first evidence for the direct regulation of PLCϵ by Gβγ and provide insights into the mechanism by which Gβγ subunits activate PLCϵ downstream of ET-1AR in cardiac hypertrophy.

Subjects/Keywords: Phosphatidylinositol signaling; Phospholipase C; Ras homolog gene family; Member A (RhoA)

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APA (6th Edition):

Madukwe, J. C. (2020). Insights into G Protein βγ Regulation of Phospholipase Cε. (Doctoral Dissertation). University of Rochester. Retrieved from http://hdl.handle.net/1802/35804

Chicago Manual of Style (16th Edition):

Madukwe, Jerry Chukwuemeka. “Insights into G Protein βγ Regulation of Phospholipase Cε.” 2020. Doctoral Dissertation, University of Rochester. Accessed September 26, 2020. http://hdl.handle.net/1802/35804.

MLA Handbook (7th Edition):

Madukwe, Jerry Chukwuemeka. “Insights into G Protein βγ Regulation of Phospholipase Cε.” 2020. Web. 26 Sep 2020.

Vancouver:

Madukwe JC. Insights into G Protein βγ Regulation of Phospholipase Cε. [Internet] [Doctoral dissertation]. University of Rochester; 2020. [cited 2020 Sep 26]. Available from: http://hdl.handle.net/1802/35804.

Council of Science Editors:

Madukwe JC. Insights into G Protein βγ Regulation of Phospholipase Cε. [Doctoral Dissertation]. University of Rochester; 2020. Available from: http://hdl.handle.net/1802/35804


Vrije Universiteit Amsterdam

2. Koppel, E.A. The in vivo function of mSIGNR1, a DC-SIGN homolog .

Degree: 2006, Vrije Universiteit Amsterdam

Subjects/Keywords: The in vivo function of mSIGNRI, a DC-SIGN homolog; mSIGNR1; host-pathogen interactions; DC-SIGN; spleen

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APA · Chicago · MLA · Vancouver · CSE | Export to Zotero / EndNote / Reference Manager

APA (6th Edition):

Koppel, E. A. (2006). The in vivo function of mSIGNR1, a DC-SIGN homolog . (Doctoral Dissertation). Vrije Universiteit Amsterdam. Retrieved from http://hdl.handle.net/1871/9884

Chicago Manual of Style (16th Edition):

Koppel, E A. “The in vivo function of mSIGNR1, a DC-SIGN homolog .” 2006. Doctoral Dissertation, Vrije Universiteit Amsterdam. Accessed September 26, 2020. http://hdl.handle.net/1871/9884.

MLA Handbook (7th Edition):

Koppel, E A. “The in vivo function of mSIGNR1, a DC-SIGN homolog .” 2006. Web. 26 Sep 2020.

Vancouver:

Koppel EA. The in vivo function of mSIGNR1, a DC-SIGN homolog . [Internet] [Doctoral dissertation]. Vrije Universiteit Amsterdam; 2006. [cited 2020 Sep 26]. Available from: http://hdl.handle.net/1871/9884.

Council of Science Editors:

Koppel EA. The in vivo function of mSIGNR1, a DC-SIGN homolog . [Doctoral Dissertation]. Vrije Universiteit Amsterdam; 2006. Available from: http://hdl.handle.net/1871/9884

3. Jorge, Uana Maria Miguel. Tumores gástricos primários múltiplos e únicos: análise imunohistoquímica comparativa.

Degree: Mestrado, Cirurgia do Aparelho Digestivo, 2006, University of São Paulo

Introdução: Adenocarcinomas gástricos múltiplos primários (AGMP) são encontrados em 3,5% a 10% de todos os pacientes com câncer gástrico. A multiplicidade tumoral é amplamente reconhecida como indicador de predisposição genética para o desenvolvimento de neoplasias Além disso, as rotas de carcinogênese não estão claramente definidas nestes tumores (rota mutadora, ou supressora, ou da E-caderina). Objetivo: avaliar a imunoexpressão de hMLH1, hMSH2, e hMSH6 (rota mutadora), p53 (rota supressora) e E-caderina nos AGMP comparando-se com adenocarcinomas únicos (pareados quanto ao sexo, idade, tipo histológico, localização e estádio) e sua relação com dados clínico-patológicos. Casuística: dezenove pacientes com AGMP foram comparados a 21 pacientes com tumores gástricos únicos quanto a características imunohistoquímicas. Métodos: Blocos de tecido fixados em formalina a 10% e incluídos em parafina foram submetidos a cortes histológicos de 4 mm, para as avaliações histológica e imunohistoquímica para hMLH1, hMSH2, hMSH6, p53 e E-caderina. Resultados: A média de idade dos pacientes com AGPM foi de 66 + 9,06 anos, e de 60 + 16,9 anos nos pacientes com tumor único (P=0,56). Vinte e dois tumores estavam localizados na porção distal do estômago; 14, no corpo e cinco na porção proximal. Em 14 pacientes, as lesões eram próximas (< 3 cm), enquanto que, em cinco pacientes, as lesões estavam em outra porção do estômago. O estágio final anatomopatológico pós-operatório foi: 15 no estágio T1 (37,5%) (8 múltiplos e 7 únicos), 7 no estágio T2 (17,5%) (1 múltiplo e 6 únicos), 17 no estágio T3 (42,5%) (9 múltiplos e 8 únicos) e 1 no estágio T4 (27,5%) (1 múltiplo). Segundo a classificação de Laurén, 45 dos tumores foram do tipo intestinal (29 múltiplos e 16 únicos), 16 do tipo gástrico (12 múltiplos e 4 únicos) e um tumor do tipo misto (1 único). O estádio anatomopatológico revelou 30 tumores avançados (16 múltiplos e 14 únicos) e 32 precoces (25 múltiplos e 7 únicos). Na imunohistoquímica, não houve diferença entre a imunoexpressão nos dois grupos de tumores quanto a: hMLH1 (24,3% vs. 19% P=0,64), hMSH6 (4,8% vs. 2,4%, P=0,68), p53 (39% vs. 24%, P=0,35) e E-caderina (27% vs. 19%, P=0,46). hMSH2 foi positivo em todos os casos. Não houve associação entre os imuno-marcadores e os dados clínico-patológicos. Conclusões: 1. As rotas de carcinogênese, mutatora, supressora e E-caderina parecem estar independentemente envolvidas no desenvolvimento dos AGMP; 2. Não houve diferença de imunoexpressão dos marcadores analisados quando compararam-se os AGMP e os tumores únicos.

Introduction: Multiple primary gastric adenocarcinomas (MPGA) have been reported from 3.5% to 10% of all patients with gastric cancer. Tumoral multiplicity is largely known as an indicator of genetic predisposition for the development of neoplasias. Moreover, the route of carcinogenesis has not been clearly clarified in these tumors (mutator pathway or suppressor pathway). Aim: to evaluate the immunoexpression of hMLH1, hMSH2, and hMSH6 (mutator pathway), p53 (suppressor…

Advisors/Committee Members: Ribeiro Júnior, Ulysses.

Subjects/Keywords: Adenocarcima; Adenocarcinoma; Caderinas; Cadherins; Genomic instability; Immunohistochemistry; Imunohistoquímica; Instabilidade genômica; Multiple primary neoplasms; Muts homolog 2 protein; Neoplasia gástricas; Neoplasias primárias múltiplas; Proteína 2 homóloga a MutS; Proteína supressora de tumor p53; Stomach neoplasms; Tumor suppressor protein p53

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APA · Chicago · MLA · Vancouver · CSE | Export to Zotero / EndNote / Reference Manager

APA (6th Edition):

Jorge, U. M. M. (2006). Tumores gástricos primários múltiplos e únicos: análise imunohistoquímica comparativa. (Masters Thesis). University of São Paulo. Retrieved from http://www.teses.usp.br/teses/disponiveis/5/5154/tde-29012007-154954/ ;

Chicago Manual of Style (16th Edition):

Jorge, Uana Maria Miguel. “Tumores gástricos primários múltiplos e únicos: análise imunohistoquímica comparativa.” 2006. Masters Thesis, University of São Paulo. Accessed September 26, 2020. http://www.teses.usp.br/teses/disponiveis/5/5154/tde-29012007-154954/ ;.

MLA Handbook (7th Edition):

Jorge, Uana Maria Miguel. “Tumores gástricos primários múltiplos e únicos: análise imunohistoquímica comparativa.” 2006. Web. 26 Sep 2020.

Vancouver:

Jorge UMM. Tumores gástricos primários múltiplos e únicos: análise imunohistoquímica comparativa. [Internet] [Masters thesis]. University of São Paulo; 2006. [cited 2020 Sep 26]. Available from: http://www.teses.usp.br/teses/disponiveis/5/5154/tde-29012007-154954/ ;.

Council of Science Editors:

Jorge UMM. Tumores gástricos primários múltiplos e únicos: análise imunohistoquímica comparativa. [Masters Thesis]. University of São Paulo; 2006. Available from: http://www.teses.usp.br/teses/disponiveis/5/5154/tde-29012007-154954/ ;

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