Naslov
IqgC at the crossroads of RasGAP and IQGAP protein families

Autori
Mijanović, Lucija ; Putar, Darija ; Filić, Vedrana ; Weber, Igor

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
4th Croatian Microscopy Congress with International Participation : Book of Abstracts / Macan, Jelena ; Kovačević, Goran - Zagreb : Hrvatsko mikroskopijsko društvo ; Institut Ruđer Bošković, 2022, 76-77

ISBN
978-953-7941-41-3

Skup
4th Croatian Microscopy Congress (CMC 2022)

Mjesto i datum
Poreč, Hrvatska, 18.05.2022. - 20.05.2022

Vrsta sudjelovanja
Poster

Vrsta recenzije
Domaća recenzija

Ključne riječi
IqgC ; Dictyostelium discoideum ; adhesion ; GAP ; IQGAP

Sažetak
Ras-specific GTPase activating proteins (RasGAPs) constitute a diverse group of proteins characterized by their GAP domain. This domain binds and inactivates small GTPases from the Ras family by stimulating their GTP hydrolytic activity. Despite having a highly homologous GAP- related domain (GRD), members of the IQ-motif containing Ras GTPase-activating-like protein (IQGAP) family do not function as RasGAPs. They are multidomain proteins that serve as scaffolds for various pathways and modulate diverse cellular processes [1]. Amoeba Dictyostelium discoideum encodes four IQGAP-related proteins – DGAP1, GAPA, IqgC and IqgD. DGAP1 and GAPA are extensively studied and exhibit traditional IQGAP activity, i.e. they participate in the formation of large protein complexes involved in the regulation of actin cytoskeleton and are unable to inactivate Ras GTPases. We showed recently that IqgC, despite apparently belonging to the IQGAP family, is a genuine RasGAP [2]. It binds and inactivates small GTPase RasG, acting as a negative regulator of large-scale endocytosis. Based on the observation that iqgC-null cells detach easily from the cell culture dishes, we set out to characterize the role of IqgC in the cell-substratum adhesion. Shaking assays showed that iqgC-null cells adhere considerably more weakly to the glass surface than the wild-type cells, and expression of recombinant IqgC in mutant cells rescued this phenotype. IqgC localizes to the punctate adhesion structures together with the adhesion marker paxillin B, as shown by total internal reflection fluorescence (TIRF) microscopy. We expressed fluorescently labeled IqgC in rasG-null cells and examined them using confocal microscopy. Normal localization of IqgC to the adhesion foci was observed, indicating that RasG is not necessary for its role in adhesion. To further dissect the role of IqgC in the cell-substratum adhesion, we expressed its individual fluorescently labeled domains in wild- type cells and examined their localization: the GRD domain with GAP activity towards RasG, and the RGCt domain unique to IQGAPs. YFP-RGCt, but not YFP-GRD, localized to the adhesion foci. The same constructs were tested for their ability to rescue the adhesion phenotype of iqgC-null cells in shaking assays, but neither could completely rescue the defect. We conclude that the interaction of IqgC with RasG and its GAP activity is not essential for the role of IqgC in the cell- substratum adhesion. The RGCt domain of IqgC is necessary and sufficient for the proper localization of the protein to the adhesion foci, but the expression of the full-length protein is necessary to restore normal adhesion. Since the RGCt domain is a hallmark of IQGAPs, our results suggest that IqgC unifies the features and activities of both RasGAP and IQGAP protein families in the regulation of various cellular processes.

Izvorni jezik
Engleski

Znanstvena područja
Biologija

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