Identifikacija fosfataze odgovorne za defosforilaciju mitohondrijskog receptora NIX

master's thesis

• Title (english): Identification of phosphatase responsible for dephosphorylation of mitophagy receptor NIX
• Author: Jadranka Škorput
• Mentor: Ivana Novak Nakir (mentor)
• Mentor: Martina Šeruga Musić (sumentor)
• Committee member: Martina Šeruga Musić (predsjednik povjerenstva)
• Committee member: Inga Urlić (član povjerenstva)
• Committee member: Sunčica Bosak (član povjerenstva)
• Committee member: Ana Galov (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2020-09-28, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Uklanjanje mitohondrija procesom mitofagije održava zdravu populaciju mitohondrija i pridonosi održavanju homeostaze stanice. Nedavna istraživanja ukazuju na važnost dimerizacije mitohondrijskog protein NIX. Dimerizacija proteina NIX prethodi indukciji mitofagije, a sama regulacija dimerizacije proteina nije poznata. Cilj ovog istraživanja je identificirati fosfatazu odgovornu za defosforilaciju mitohondrijskog proteina NIX i regulaciju dimerizacije. Proučavanjem znanstvene literature, odabrana je PP1 fosfataza. Interakcija PP1 fosfataze i proteina NIX, kao i njegovih mutanti, ispitana je in vitro u staničnoj liniji HEK293 pull down imunoprecipitacijom i Western blot imunodetekcijom. Utjecaj PP1 fosfataze na indukciju mitofagije, provjerio se metodom RNA interferencije in vivo u modelnom organizmu Caenorhabditis elegans u normalnim uvjetima, kao i u prisutnosti mitohondrijskog otrova – parakvat. Dobivenim rezultatima potvrđena je interakcija PP1 fosfataze i proteina NIX u staničnoj liniji HEK293 te je utvrđeno da PP1 fosfataza inhibira mitofagiju u normalnim uvjetima, ali ne i pri indukciji mitofagije.

Abstract (english)

Removal of mitochondria, by the process of mitophagy, maintains a healthy population of mitochondria and contributes to maintenance of cell homeostasis. Recent research indicates the importance of mitochondrial protein NIX dimerization, whereas the regulation of protein dimerization remains unknown. The aim of this study was to identify the phosphatase responsible for dephosphorylation of the mitochondrial protein NIX, thus regulating the dimerization of protein NIX. Phosphatase PP1 was selected by searching the literature. Pull-down and Western blot analysis were used to investigate the interaction of PP1 phosphatase with NIX protein, as well as its mutants, in vitro in HEK293 cells. The effect of PP1 phosphatase in mitophagy induction was investigated with RNA interference assay in vivo in model organism Caenorhabditis elegans, both under normal conditions, as well as in the presence of mitochondrial toxin - paraquat. The results confirm the interaction between PP1 phosphatase and mitochondrial protein NIX. Moreover, the results show that PP1 inhibits mitophagy under normal conditions but show no impact after the induction of mitophagy.

• Keywords: autofagija; mitofagija; NIX; PP1; fosfataza
• Language: croatian
• URN:NBN: urn:nbn:hr:217:753215
• Study programme: Title: Molecular Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra molekularne biologije (magistar/magistra molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access; Embargo expiration date: 2022-09-28
• Created on: 2020-10-07 09:17:55