Identification of Alzheimer’s disease relevant interacting proteins using “in silico” and cellular procedures

• Autor(a) principal: Carvalho, Estefânia Coelho
• Data de Publicação: 2019
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10773/28386
• topic: Aβ; Protein complexes; SH-SY5Y cell line; GRB2; TP53BP2; Alzheimer’s Disease

APP is a protein of relevance to Alzheimer´s disease as it is the precursor for the Aβ peptide that is the major constituent of senile plaques seen deposited in the brains of individuals with Alzheimer’s Disease. Dephosphorylation, by phosphatases like PP1, is among the most common forms of post-translational modification in proteins and anomalies in this process have been linked to Alzheimer’s disease. Therefore, it may be important to identify APP binding proteins which can be modulated by phosphatases such as PP1. Here we focus on two proteins, TP53BP2 and GRB2. TP53BP2 is a protein that plays a central role in regulating apoptosis and cell growth via its interactions with other proteins such as the members of the p53 family. GRB2 is an adapter protein that provides a connection between cell surface growth factor receptors and the Ras signaling pathway. Firstly, we show that neither of these proteins’ expression levels are affected by exposure to previously aggregated Aβ42 in undifferentiated SH-SY5Y cells. Secondly, relying on yeast two hybrid assays previously carried out by our research group using PP1 or APP as bait, we hypothesized that TP53BP2 and GRB2 could independently be the bridging protein in a trimeric complex involving APP and PP1. After performing co-Immunoprecipitations we were able to validate the interactions between TP53BP2 and PP1 and between GRB2 and APP, while the interactions between TP53BP2 and APP and between GRB2 and PP1 were not detected.