Uloga proteina IqgC u regulaciji makropinocitoze u amebi Dictyostelium discoideum

doctoral thesis

• Title (english): The role of IqgC protein in the regulation of macropinocytosis in amoeba Dictyostelium discoideum
• Author: Darija Putar Brajković
• Mentor: Vedrana Filić Mileta (mentor)
• Committee member: Maja Herak-Bosnar (predsjednik povjerenstva)
• Committee member: Igor Weber (član povjerenstva)
• Committee member: Biljana Balen (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; Zagreb
• Defense date and country: 2024-02-21, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology
• Universal decimal classification (UDC): 577 - Biochemistry. Molecular biology. Biophysics

Abstract

Protein IqgC iz protista D. discoideum djeluje kao GAP (GTPase activating protein) prema GTPazi RasG, čime je deaktivira i tako suprimira makropinocitozu. Međutim, kako je makropinocitoza u akseničnim sojevima ovog organizma konstitutivno pojačana, delecija gena iqgC u akseničnom soju rezultirala je neznatnim pojačanjem makropinocitoze. U ovom radu pokazala sam da delecija gena iqgC ima znatno veću pozitivnu ulogu u regulaciji makropinocitoze u soju divljeg tipa čija makropinocitoza je uvelike suprimirana. Nadalje, ustanovila sam da je RasG neophodan za vezanje IqgC na membranu makropinosoma, dok interakcija IqgC s fosfoinozitidima doprinosi stabilizaciji vezanja. Pokazala sam također da IqgC izravno interagira s GTPazom Rab5A, ali ne utječe na njenu GTPaznu aktivnost, dok suvišak Rab5A inhibira GAP aktivnost IqgC prema RasG in vitro. Također, prekomjerna ekspresija Rab5A izaziva preuranjenu disocijaciju IqgC s makropinosoma. Dobiveni rezultati pokazuju da, osim interakcije proteina IqgC i RasG, interakcija IqgC i Rab5A također igra ulogu u regulaciji makropinocitoze.

Abstract (english)

The D. discoideum protein IqgC is a RasG-specific GAP (GTPase activating protein) that negatively regulates macropinocytosis. However, deletion of iqgC in the axenic strain induced only a minor enhancement of macropinocytosis, as axenic cells already exhibit upregulated fluid uptake. Therefore, in this work, I deleted iqgC in the wild-type strain, whose macropinocytosis is largely suppressed, and demonstrated a more pronounced positive effect on fluid uptake upon IqgC depletion. Furthermore, I demonstrated that RasG is essential for IqgC recruitment to macropinosomes, while IqgC binding to phosphoinositides likely contributes to stabilization of the association. I showed that IqgC also interacts with GTPase Rab5A but does not affect its activity. Actually, an excessive amount of Rab5A inhibits the GAP activity of IqgC towards RasG in vitro and induces premature dissociation of IqgC from the macropinosome in vivo. These results suggest that interactions between IqgC and both GTPases play a regulatory role in macropinocytosis.

• Keywords: IqgC; RasG; Rab5; RasGAP; makropinocitoza; Dictyostelium
• Keywords (english): IqgC; RasG; Rab5; RasGAP; macropinocytosis; Dictyostelium
• Language: croatian
• URN:NBN: urn:nbn:hr:217:491160
• Promotion: 2024
• Project: Number: IP-2020-02-1572; Title: Regulacija endocitoze na velikoj skali pomoću IQGAP proteinima srodnih proteina IqgC i IqgD; Acronym: RegEndIqCD; Leader: Vedrana Filić Mileta; Jurisdiction: Croatia; Funder: HRZZ; Funding stream: IP
• Project: Number: 180584; Title: Phagocytosis and Macropinocytosis, a mechanistic view; Leader: Igor Weber; Jurisdiction: Croatia; Funder: HRZZ; Funding stream: HSIP
• Study programme: Title: Doctoral study; Study programme type: university; Study level: postgraduate; Academic / professional title: doktor/doktorica znanosti u području prirodnih znanosti (doktor/doktorica znanosti u području prirodnih znanosti)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2024-02-27 10:34:53