dc.contributor.author
Mostafa Hassan Elkewedi, Mohamed
dc.date.accessioned
2019-04-03T12:04:09Z
dc.date.available
2019-04-03T12:04:09Z
dc.identifier.uri
https://refubium.fu-berlin.de/handle/fub188/24287
dc.identifier.uri
http://dx.doi.org/10.17169/refubium-2059
dc.description.abstract
Glycogen synthase kinase 3β interaction protein (GSKIP), a ubiquitously expressed protein, was identified as a direct interaction partner of glycogen synthase kinase 3β (GSK3β) and protein kinase A (PKA). It was found to facilitate the PKA-mediated inhibitory phosphorylation of GSK3β, as well as modulate the phosphorylation of GSK3β and PKA substrates. Through this modulation, GSKIP has been implicated in the fine tuning of canonical Wnt signalling and was found to play a role in both mitochondrial fission and cancer cells dynamics. Building upon these findings, this thesis uncovers novel roles for GSKIP in actin cytoskeleton reorganization and maintenance of the epithelial phenotype.
The reorganization of the cellular actin cytoskeleton is essential for various processes, among which are cell signalling, and the development and maintenance of cellular junctions and polarity. The biphasic modulation of actin dynamics is attained by the actin severing protein cofilin (CFL), which can induce either the disassembly or the assembly of the actin filaments through its ability to depolymerize and sever the filaments. The activity of CFL is negatively regulated by the Rho GTPases, p38 mitogen-activated protein kinase (p38 MAPK),
and PKA mediated phosphorylation at S3. The knockdown (KD) of GSKIP in various cancer cells was found to modulate this phosphorylation, with the human non-small lung adenocarcinoma cells, A549 and the human cervical adenocarcinoma cells, HeLa-S3 exhibiting the most marked decrease in phosphorylation. The KD of GSKIP in A549 cells was characterized by increased actin depolymerization at the cellular junctions and decreased CFL abundance in the filamentous polymerized actin fraction. Similarly, the KD of GSKIP in HeLa-S3 cells was associated with phenotypic changes and an altered actin cytoskeleton, which was marked by increased actin stress fiber formation. The molecular mechanisms underlying these cytoskeletal changes were found to be most likely PKA-dependent, since the KD of GSKIP in the studied cells did not alter either the Rho GTPases or the p38 MAPK-mediated phosphorylation of CFL. Taken together, these findings implicate GSKIP as a modulator of the PKA-mediated CFL-regulated actin dynamics.
Investigating the GSKIP KD-induced phenotypic changes revealed a shift of the cells from the epithelial to the mesenchymal phenotype. Studying the associated molecular mechanisms showed downregulation of prominent epithelial markers, such as E-cadherin and β-catenin, as well as upregulation of the epithelial to mesenchymal transition (EMT) inducer ZEB1 in A549 and HeLa-S3 cells. The GSKIP KD-mediated loss of the epithelial phenotype was confirmed by investigating the integrity of crucial epithelial junctions, namely the adherens junction and the desmosomes. Both junctions displayed marked anomalies upon the KD of GSKIP, suggesting a decrease in the integrity of both and uncovering the essentiality of GSKIP for the maintenance of epithelial junctions.
en
dc.format.extent
114 Seiten
dc.rights.uri
http://www.fu-berlin.de/sites/refubium/rechtliches/Nutzungsbedingungen
dc.subject
PKA Signalling
en
dc.subject.ddc
500 Natural sciences and mathematics::500 Natural sciences::500 Natural sciences and mathematics
dc.title
Elucidation of Functions of the AKAP GSKIP
dc.contributor.gender
male
dc.contributor.firstReferee
Klußmann, Enno
dc.contributor.furtherReferee
Freund, Christian
dc.date.accepted
2019-03-19
dc.identifier.urn
urn:nbn:de:kobv:188-refubium-24287-3
refubium.affiliation
Biologie, Chemie, Pharmazie
dcterms.accessRights.dnb
free
dcterms.accessRights.openaire
open access
dcterms.accessRights.proquest
accept