doktorska disertacija
Katja Leben (Author), Roman Jerala (Mentor)

Abstract

Narava uravnava številne celične procese z vplivom na izražanje genov. Prepisovanje DNA v evkariontskih celicah sproži medsebojno delovanje pestrega nabora proteinov. Poznavanje njihovega kooperativnega delovanja je pomembno za načrtovanje uravnavanja celičnih procesov v sintezni biologiji. Razumevanje tega procesa odpira možnosti načrtovanja novih regulatornih vezij. V doktorskem delu smo preiskovali vpliv efektorja, podobnega transkripcijskemu aktivatorju (TALE), ki poveča aktivnost transkripcijskega dejavnika, vezanega tudi več deset baznih parov stran. Uporabljeni proteini TALE niso vsebovali lastne aktivacijske domene in so ojačali delovanje drugega transkripcijskega dejavnika ne glede na orientacijo, položaj navzgor ali navzdol od vezavnega mesta transkripcijskega dejavnika in razmik med vezavnima mestoma. Vpliv proteina TALE na transkripcijo smo demonstrirali na različnih sesalskih celičnih linijah, v kombinaciji s tremi različnimi DNA vezavnimi domenami, tako za aktivatorje kot represorje transkripcije. Določili smo najkrajšo razdaljo pri kateri se pojavi vpliv na drugi transkripcijski dejavnik in okarakterizirali razpon razdalj med vezavnima mestoma, kjer je vpliv še opazen. V prvem delu doktorata smo predstavili sistem za preverjanje sinergističnega delovanja proteina TALE z več različnimi DNA vezavnimi proteini. Predvidevamo, da bi lahko preko tega učinka izboljšali razumevanje kooperativnega delovanja in vloge DNA pri uravnavanju izražanja genov. Odkritje, da proteini TALE povečajo uravnavanje transkripcije z drugim transkripcijskim dejavnikom, širi raznolikost načinov na katere lahko proteini TALE vplivajo na transkripcijo in bi lahko delovali kot dodatni modulator pri uravnavanju transkripcijskih poti. Proteini s poznanim kooperativnim delovanjem predstavljajo zanimiv potencial v kontekstu sintezne biologije. Poleg tega poznavanje vpliva TALE na učinkovitost drugih transkripcijskih dejavnikov predstavlja DNA vezavni protein z okarakteriziranimi kooperativnimi lastnostmi in orodje za nadaljnje preiskovanje mehanizmov medsebojnega delovanja proteinov pri uravnavanju transkripcije in izkoriščanje njihovega potenciala.

Keywords

biokemija;DNA;nukleotidi;transkripcija;DNA-vezavne beljakovine;doktorske disertacije;Molekularna biologija;Disertacije;Sintezna biologija;Izražanje genov;

Data

Language: Slovenian
Year of publishing:
Typology: 2.08 - Doctoral Dissertation
Organization: UL MF - Faculty of Medicine
Publisher: [K. Leben]
UDC: 577.2(043.3)
COBISS: 134072835 Link will open in a new window
Views: 140
Downloads: 27
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Other data

Secondary language: English
Secondary title: Transcriptional regulation design based on the effect of multiple DNA-binding proteins
Secondary abstract: Several cellular processes in nature are directly dictated by gene expression. Transcription in eukaryotic cells is regulated through cooperative and synergistic actions of an ensemble of proteins, and deciphering the cooperative mechanisms could be used as a new method to design the regulation of cellular processes. Understanding this process would enable the design of intricate regulatory circuits in synthetic biology. This dissertation focuses on the effect transcription activator like effector (TALE) has on the activity of another transcription factor bound tens of base pairs away. The TALE protein, which lacks an activation domain, can enhance the effect of another transcription factor regardless of its orientation, its position upstream or downstream of the transcription factor binding site, and the distance between the respective binding sites. The effect of the TALE protein was demonstrated in several mammalian cell lines in combination with three structurally diverse DNA binding domains, regardless of whether they activate or repress transcription. The minimum and maximum distance between binding sites where the effect of TALE protein could be observed were determined. The first part of the dissertation describes a platform for investigating a potential synergistic effect of the TALE protein with diverse transcription factors, and providing mechanistic insights into cooperative interactions and the role of DNA in translation regulation. The discovered ability of TALE proteins to enhance transcriptional regulation of other transcription factors further expands the variety of the effects of TALE proteins on transcription, which could act as an additional modulator in regulating transcriptional pathways. Allosteric proteins have an interesting potential as tools for synthetic biology. In addition to expanding our understanding of the variety of synergistic effects on transcriptional regulation, a designable DNA binding protein with characterized cooperative properties could serve as a tool to investigate the mechanisms of synergistic transcriptional regulation and harvest its potential.
Secondary keywords: Molekularna biologija;Univerzitetna in visokošolska dela;
Type (COBISS): Doctoral dissertation
Study programme: 0
Embargo end date (OpenAIRE): 1970-01-01
Thesis comment: Univ. v Ljubljani, Medicinska fak.
Pages: VII, 44, IX f., 13 f. pril.
ID: 16765068