Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease

Detalhes bibliográficos
Autor(a) principal: Correia, Susana Margarida da Silva
Data de Publicação: 2015
Tipo de documento: Dissertação
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10773/15769
Resumo: The present study was undertaken to identify proteins interacting with PrPC that could provide new insights into its physiological functions and pathological role. We performed a target search for lysosomal network protein, Rab7a and Rab9, in frontal cortex and cerebellum of human brain from patients with sCJD-MM1 and sCJD-VV2. The intracellular level of Rab7a was increased significantly, when compared with healthy age-matched control. Interactions of PrPC and Rab7a/Rab9 were further investigated by using confocal laser scanning microscopy. Immunofluorescence results suggested potential interactions of Rab7a and PrPC. siRNA against the Rab7a gene was used to knockdown the expression of Rab7a protein in primary cell culture of cortical neurons from wild type mice. This depleted Rab7a resulted an impairment of PrPC trafficking leading to an accumulation of PrPC in the endocytosis pathway. Furthermore, interactions of Tau and Rab7a were investigated by using western blot analysis and confocal laser scanning microscopy. Cell cultures of cortex of wildtype mice were treated with siRNA-Tau, siRNA-Rab7 and control siRNA followed by immunofluorescence. The results of immunofluorescence suggested potential interaction of Tau and Rab7a. Cells lines treated with siRNA-Tau, the intracellular levels of Rab7a and Rab9 significantly increases and their localization is also modified. When we transfected this cells lines with siRNA-rab7a the accumulation of Tau decreases in cytosolic region and their localization was also modified when compared with control cells. In conclusion, this study may help to understand and characterize the subtype specific disease progression in CJD cases. Furthermore, it could be a step ahead to development of new treatment strategies for diseases subtype specific manner.
id RCAP_b43fd00b96d1ea0a0a02a4d8a22bf29a
oai_identifier_str oai:ria.ua.pt:10773/15769
network_acronym_str RCAP
network_name_str Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
repository_id_str 7160
spelling Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob diseaseBiologia molecular e celularDoença de Creutzfeldt-JakobProteína precursora de amilóidePriões - DoençasThe present study was undertaken to identify proteins interacting with PrPC that could provide new insights into its physiological functions and pathological role. We performed a target search for lysosomal network protein, Rab7a and Rab9, in frontal cortex and cerebellum of human brain from patients with sCJD-MM1 and sCJD-VV2. The intracellular level of Rab7a was increased significantly, when compared with healthy age-matched control. Interactions of PrPC and Rab7a/Rab9 were further investigated by using confocal laser scanning microscopy. Immunofluorescence results suggested potential interactions of Rab7a and PrPC. siRNA against the Rab7a gene was used to knockdown the expression of Rab7a protein in primary cell culture of cortical neurons from wild type mice. This depleted Rab7a resulted an impairment of PrPC trafficking leading to an accumulation of PrPC in the endocytosis pathway. Furthermore, interactions of Tau and Rab7a were investigated by using western blot analysis and confocal laser scanning microscopy. Cell cultures of cortex of wildtype mice were treated with siRNA-Tau, siRNA-Rab7 and control siRNA followed by immunofluorescence. The results of immunofluorescence suggested potential interaction of Tau and Rab7a. Cells lines treated with siRNA-Tau, the intracellular levels of Rab7a and Rab9 significantly increases and their localization is also modified. When we transfected this cells lines with siRNA-rab7a the accumulation of Tau decreases in cytosolic region and their localization was also modified when compared with control cells. In conclusion, this study may help to understand and characterize the subtype specific disease progression in CJD cases. Furthermore, it could be a step ahead to development of new treatment strategies for diseases subtype specific manner.O presente estudo foi levado a cabo com o intuito de identificar possíveis proteínas que interajam com PrPC que possam fornecer novos conhecimentos sobre as suas funções fisiológicas assim como no papel patológico. O presente estudo incidiu na investigação dos níveis das proteínas endossomais Rab7a e Rab9 em amostras do córtex frontal e cerebelo de pacientes diagnosticados com sCJD-MM1 e sCJD-VV2. Observamos um aumento significativo dos níveis intracelulares do Rab7a em pacientes diagnosticados com sCJD-MM1 e sCJD-VV2 quando comparadas com amostras de pacientes na mesma faixa etária sem a doença (controlo). As interações entre PrPC e as proteínas Rab7a e Rab9 foram posteriormente estudadas com o auxílio à microscopia confocal. Os resultados da imunofluorescência sugeriram potenciais interações entre as proteínas Rab7a com o PrPC. O siRNA-Rab7 foi usado para diminuir a expressão da proteína Rab7a (“Knockdown”) na cultura primária de células do córtex de ratos saudáveis. Na cultura de células do córtex tratadas com o siRNA-Rab7, foi observado um emparelhamento da via endocítica. Seguidamente investigamos possíveis interações da proteína Tau com a proteína Rab7a, recorrendo ao western blot e a microscopia confocal. Culturas de células do córtex de ratos saudáveis foram tratadas com siRNA-Tau, siRNA-Rab7a e siRNA. Os resultados da imunofluorescência das diferentes culturas celulares sugeriram uma potencial interação entre as proteínas Tau e Rab7a. Nas linhas celulares tratadas com siRNA-Tau os níveis intracelulares das proteínas Rab7a e Rab9 aumentaram significativamente, assim como, foi também observada a alteração da sua localização. Nas células tratadas com siRNA-Rab7a observamos uma diminuição da acumulação da proteína Tau na região citosólica. Em conclusão, este trabalho pode ajudar a perceber e a caracterizar a progressão da doença nos subtipos específicos, no caso de sCJD. Contudo, este trabalho poderá ser também um passo à frente para o desenvolvimento de novas estratégias terapêuticas para os subtipos sCJD-MM1 e sCJD-VV2.Universidade de Aveiro2016-06-20T14:44:44Z2015-01-01T00:00:00Z2015info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisapplication/pdfhttp://hdl.handle.net/10773/15769engCorreia, Susana Margarida da Silvainfo:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2024-02-22T11:29:15Zoai:ria.ua.pt:10773/15769Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T02:51:04.224207Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse
dc.title.none.fl_str_mv Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
title Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
spellingShingle Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
Correia, Susana Margarida da Silva
Biologia molecular e celular
Doença de Creutzfeldt-Jakob
Proteína precursora de amilóide
Priões - Doenças
title_short Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
title_full Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
title_fullStr Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
title_full_unstemmed Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
title_sort Altered regulatory response of Rab7a and Rab9 in MM1 and VV2 subtype of Creutzfeldt-Jakob disease
author Correia, Susana Margarida da Silva
author_facet Correia, Susana Margarida da Silva
author_role author
dc.contributor.author.fl_str_mv Correia, Susana Margarida da Silva
dc.subject.por.fl_str_mv Biologia molecular e celular
Doença de Creutzfeldt-Jakob
Proteína precursora de amilóide
Priões - Doenças
topic Biologia molecular e celular
Doença de Creutzfeldt-Jakob
Proteína precursora de amilóide
Priões - Doenças
description The present study was undertaken to identify proteins interacting with PrPC that could provide new insights into its physiological functions and pathological role. We performed a target search for lysosomal network protein, Rab7a and Rab9, in frontal cortex and cerebellum of human brain from patients with sCJD-MM1 and sCJD-VV2. The intracellular level of Rab7a was increased significantly, when compared with healthy age-matched control. Interactions of PrPC and Rab7a/Rab9 were further investigated by using confocal laser scanning microscopy. Immunofluorescence results suggested potential interactions of Rab7a and PrPC. siRNA against the Rab7a gene was used to knockdown the expression of Rab7a protein in primary cell culture of cortical neurons from wild type mice. This depleted Rab7a resulted an impairment of PrPC trafficking leading to an accumulation of PrPC in the endocytosis pathway. Furthermore, interactions of Tau and Rab7a were investigated by using western blot analysis and confocal laser scanning microscopy. Cell cultures of cortex of wildtype mice were treated with siRNA-Tau, siRNA-Rab7 and control siRNA followed by immunofluorescence. The results of immunofluorescence suggested potential interaction of Tau and Rab7a. Cells lines treated with siRNA-Tau, the intracellular levels of Rab7a and Rab9 significantly increases and their localization is also modified. When we transfected this cells lines with siRNA-rab7a the accumulation of Tau decreases in cytosolic region and their localization was also modified when compared with control cells. In conclusion, this study may help to understand and characterize the subtype specific disease progression in CJD cases. Furthermore, it could be a step ahead to development of new treatment strategies for diseases subtype specific manner.
publishDate 2015
dc.date.none.fl_str_mv 2015-01-01T00:00:00Z
2015
2016-06-20T14:44:44Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/masterThesis
format masterThesis
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://hdl.handle.net/10773/15769
url http://hdl.handle.net/10773/15769
dc.language.iso.fl_str_mv eng
language eng
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv Universidade de Aveiro
publisher.none.fl_str_mv Universidade de Aveiro
dc.source.none.fl_str_mv reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
instacron:RCAAP
instname_str Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
instacron_str RCAAP
institution RCAAP
reponame_str Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
collection Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
repository.name.fl_str_mv Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
repository.mail.fl_str_mv
_version_ 1799137560176361472