Utjecaj transmembranskog proteina AQP5 na ekspresiju proteina FOXO1, NRF2 i AQP3 u ljudskim stanicama tumora dojke

master's thesis

• Title (english): The effect of AQP5 transmembrane protein on FOXO1, NRF2 and AQP3 expression in human breast tumor cells
• Author: Iva Žulj
• Mentor: Ana Čipak Gašparović (mentor)
• Mentor: Maja Matulić (komentor)
• Committee member: Maja Matulić (predsjednik povjerenstva)
• Committee member: Anamaria Štambuk (član povjerenstva)
• Committee member: Sofia Ana Blažević (član povjerenstva)
• Committee member: Petra Peharec Štefanić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2023-12-19, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Akvaporini su membranske pore kojima prolaze voda, glicerol i druge male molekule u stanicu i iz nje, čime sudjeluju u regulaciji važnih staničnih funkcija. Sukladno tome, povećana ekspresija akvaporina 3 (AQP3) i akvaporina 5 (AQP5) korelira s malignošću tumora dojke. AQP3 i AQP5 pripadaju posebnoj skupini akvaporina, peroksiporinima, koji prenose H2O2. Ulaskom u stanicu H2O2 utječe na signalne puteve, a u većim koncentracijama uzrokuje oksidacijski stres. Važnu ulogu u odgovoru stanice na oksidacijski stres imaju transkripcijski faktori FOXO1 i NRF2 koji reguliraju transkripciju antioksidacijskih enzima poput katalaze i glutation sintetaze. Stoga je cilj ovog istraživanja bio utvrditi utječe li prekomjerna ekspresija AQP5 na transkripcijske faktore FOXO1 i NRF2 i na aktivnost njihovih ciljnih gena. Istraživanje je provedeno na trima staničnim linijama tumora dojke: MCF7 (pozitivna na receptore estrogena i progesterona), SkBr3 (pozitivna na receptor HER2) i SUM159 (negativna na receptore hormona i HER2) i iz njih izvedenih linija, uzgojenih nakon transfekcije plazmidom pCMV6 i plazmidom sa sekvencom koja kodira AQP5. Metodama Western blota i PCR-a praćen je utjecaj povećane ekspresije AQP5 na ekspresiju gena i proteina FOXO1, NRF2 i AQP3. Određene su koncentracije glutationa i aktivnost katalaze. Rezultati ukazuju na moguću povezanost prekomjerne ekspresije AQP5 s povećanjem ekspresije AQP3 te s FOXO1.

Abstract (english)

Aquaporins are membrane pores that facilitate the transport of water, glycerol and other small molecules across the cell membrane and are therefore involved in the regulation of important cellular functions. Accordingly, the upregulation of aquaporin 3 (AQP3) and aquaporin 5 (AQP5) is associated with malignancy of breast cancer. AQP3 and AQP5 are members of a special group of aquaporins, peroxiporins, which facilitate H2O2 transport. Once in the cell, H2O2 affects different signaling pathways, while at higher concentrations it induces oxidative stress. FOXO1 and NRF2 are transcription factors that play pivotal role in cellular response to oxidative stress and regulate the transcription of antioxidant enzymes such as catalase and glutathione synthetase. Hence, the aim of this study was to investigate whether the overexpression of AQP5 affects the transcription factors FOXO1 and NRF2 and the activity of their target genes. Three breast cancer cell lines were used in this study: MCF7 (positive for estrogen and progesterone receptors), SkBr3 (HER2 positive) and SUM159 (negative for hormone receptors and HER2) as well as cell lines derived by transfection with plasmid pCMV6 and plasmid containing sequence coding for AQP5. Western blot and PCR methods were used to assess the influence of increased AQP5 expression on FOXO1, NRF2 and AQP3, on RNA and protein level. In addition, glutathione concentrations and catalase activity were measured. The results suggest a possible link between AQP5 overexpression and an increase in AQP3 expression as well as a plausible link with FOXO1.

• Keywords: akvaporini; peroksiporini; antioksidacijska zaštita; MCF7; SkBr3; SUM159
• Keywords (english): aquaporins; peroxiporins; antioxidative defense; MCF7; SkBr3; SUM159
• Language: croatian
• URN:NBN: urn:nbn:hr:217:948831
• Project: Number: IP-2020-02-3617; Title: Određivanje uloga akvaporina 3 i 5 u razvoju rezistencije tumora dojke na oksidacijski stres; Title: Elucidating the role of aquaporins 3 and 5 in the development of breast cancer resistance to oxidative stress; Acronym: AquaBCaRe; Leader: Ana Čipak Gašparović; Jurisdiction: Croatia; Funder: HRZZ; Funding stream: IP
• Study programme: Title: Molecular Biology-University graduate programme; Study programme type: university; Study level: graduate; Academic / professional title: sveučilišni/a magistar/magistra molekularne biologije (sveučilišni/a magistar/magistra molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Embargoed access; Embargo expiration date: 2025-12-19
• Created on: 2024-01-15 13:19:22