Uloga RNA helikaze DDX5 u signalnom putu NF-kB

master's thesis

• Title (english): The role of DDX5 RNA helicase in NF-kB signaling
• Author: Sandra Postić
• Mentor: Rainer de Martin (mentor)
• Mentor: Maja Matulić (sumentor)
• Committee member: Maja Matulić (predsjednik povjerenstva)
• Committee member: Sven Jelaska (član povjerenstva)
• Committee member: Tomislav Ivanković (član povjerenstva)
• Committee member: Jasna Hrenović (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2018-02-23, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

DEAD-kutija RNA helikaza DDX5 uključena je u brojne stanične procese kroz metabolizam RNA ili kao koregulator raznih transkripcijskih faktora. U ovom radu istražen je utjecaj proteina DDX5 u signalnom putu NF-кB i staničnom ciklusu. Kako bismo dobili stanice HeLa bez ekspresije DDX5 ili s delecijom C-terminalnog kraja DDX5 proteina koji sadrži proteinsku domenu Pfam za koju se smatra da je odgovorna za aktivaciju puta NFκB, korištena je metoda delecije pomoću plazmida CRISPR/Cas9. Delecija gena DDX5 bila je letalna za stanice. Stanice HeLa s delecijom dijela sekvence DDX5, HeLa DDX5ΔPfam bile su vijabilne, ali su također pokazivale smanjenu stopu umnožavanja. Usporedno su metodom unosa antisens RNA za DDX5 uzgojene stanice HeLa i primarne stanice HUVEC sa utišanom ekspresijom DDX5, te se istražio utjecaj ove promjene na NF-κB aktivaciju i stanični ciklus.

Abstract (english)

The DEAD-box RNA helicase DDX5 is included in all sort of cellular processes through its ability to metabolize RNA or as a co-regulator of different transcription factors. In this study, the function of the DDX5 protein was investigated in the context of NF-κB signaling and proliferation. The CRISPR/Cas9 genome editing method was employed to generate DDX5-gene knockout, as well as C-terminal deleted DDX5 HeLa cells, the latter comprising a pfam-protein domain assumed to be responsible for the activation of NF-κB. While complete DDX5 knockout cells were not able to proliferate, DDX5ΔPfam HeLa cells were viable, but exhibited a reduced cell growth rate. Complementary to the CRISPR/Cas9 gene editing, short-hairpin mediated RNA interference (shRNA) was employed to establish DDX5 knock- down in HeLa, as well as primary HUVEC cells and its impact on NF-κB activation and cell cycle was investigated.

• Keywords: proliferacija; stanični ciklus; CRISPR/Cas9; HUVEC; shRNA
• Keywords (english): proliferation; cell cycle; CRISPR/Cas9; HUVEC; shRNA
• Language: croatian
• URN:NBN: urn:nbn:hr:217:924642
• 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: 2020-02-23
• Created on: 2018-03-15 12:48:06