Ezrin interacts with the scaffold protein IQGAP1 and affects its cortical localization

Nammalwar, Rathangadhara Chakrapani; Heil, Annika; Gerke, Volker

doi:10.1016/j.bbamcr.2014.12.026

Cited by 10 publications

(12 citation statements)

References 25 publications

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“…IQGAP1 is a scaffolding protein with five identified protein binding domains ( White, et al, 2012 ; Nammalwar, et al, 2015 ; Nauert, et al, 2003 ; Jacquemet, et al, 2013 ; Johnson, et al, 2013 ; Hedman, et al, 2015 ). It binds and/or stabilizes ezrin, CDC42 and RAC1, and interacts with a number of cytoskeletal and cell adhesion molecules (e.g., mDia and Cadherin) ( White, et al, 2012 ; Nammalwar, et al, 2015 ; Nauert, et al, 2003 ; Jacquemet, et al, 2013 ; Johnson, et al, 2013 ; Hedman, et al, 2015 ). As such, IQGAP1 plays a role in the regulation of signal transduction, cytoskeleton, cell adhesion and cell cycle ( White, et al, 2012 ; Hedman, et al, 2015 ).…”

Section: Resultsmentioning

confidence: 99%

Identification of IQGAP1 as a SLC26A4 (Pendrin)-Binding Protein in the Kidney

Barone

Kia

et al. 2022

Front. Mol. Biosci.

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Background: Several members of the SLC26A family of transporters, including SLC26A3 (DRA), SLC26A5 (prestin), SLC26A6 (PAT-1; CFEX) and SLC26A9, form multi-protein complexes with a number of molecules (e.g., cytoskeletal proteins, anchoring or adaptor proteins, cystic fibrosis transmembrane conductance regulator, and protein kinases). These interactions provide regulatory signals for these molecules. However, the identity of proteins that interact with the Cl−/HCO3− exchanger, SLC26A4 (pendrin), have yet to be determined. The purpose of this study is to identify the protein(s) that interact with pendrin.Methods: A yeast two hybrid (Y2H) system was employed to screen a mouse kidney cDNA library using the C-terminal fragment of SLC26A4 as bait. Immunofluorescence microscopic examination of kidney sections, as well as co-immunoprecipitation assays, were performed using affinity purified antibodies and kidney protein extracts to confirm the co-localization and interaction of pendrin and the identified binding partners. Co-expression studies were carried out in cultured cells to examine the effect of binding partners on pendrin trafficking and activity.Results: The Y2H studies identified IQ motif-containing GTPase-activating protein 1 (IQGAP1) as a protein that binds to SLC26A4’s C-terminus. Co-immunoprecipitation experiments using affinity purified anti-IQGAP1 antibodies followed by western blot analysis of kidney protein eluates using pendrin-specific antibodies confirmed the interaction of pendrin and IQGAP1. Immunofluorescence microscopy studies demonstrated that IQGAP1 co-localizes with pendrin on the apical membrane of B-intercalated cells, whereas it shows basolateral expression in A-intercalated cells in the cortical collecting duct (CCD). Functional and confocal studies in HEK-293 cells, as well as confocal studies in MDCK cells, demonstrated that the co-transfection of pendrin and IQGAP1 shows strong co-localization of the two molecules on the plasma membrane along with enhanced Cl−/HCO3− exchanger activity.Conclusion: IQGAP1 was identified as a protein that binds to the C-terminus of pendrin in B-intercalated cells. IQGAP1 co-localized with pendrin on the apical membrane of B-intercalated cells. Co-expression of IQGAP1 with pendrin resulted in strong co-localization of the two molecules and increased the activity of pendrin in the plasma membrane in cultured cells. We propose that pendrin’s interaction with IQGAP1 may play a critical role in the regulation of CCD function and physiology, and that disruption of this interaction could contribute to altered pendrin trafficking and/or activity in pathophysiologic states.

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Section: Resultsmentioning

confidence: 99%

Identification of IQGAP1 as a SLC26A4 (Pendrin)-Binding Protein in the Kidney

Barone

Kia

et al. 2022

Front. Mol. Biosci.

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“…pGEX4T Moesin-FERM was a gift from Vijaya Ramesh (Addgene plasmid # 1163 [46]). pGEX4T1 Ezrin FERM domain was a kind gift from Volker Gerke [47].…”

Section: Plasmids Cloning and Mutagenesismentioning

confidence: 99%

Short linear motif based interactions and dynamics of the ezrin, radixin, moesin and merlin FERM domains

Ali

Khramushin

Yadav

et al. 2020

Preprint

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The ERMs (ezrin, radixin and moesin) and the closely related merlin (NF2) participate in signaling events at the cell cortex through interactions mediated by their conserved FERM domain. We systematically investigated the FERM domain mediated interactions with short linear motifs (SLiMs) by screening the FERM domains againsts a phage peptidome representing intrinsically disordered regions of the human proteome. We uncovered a diverse set of interacting partners with similar but distinct binding motifs (FYDF, xYxV, FY(D/E)L and LQE(I/L) that bind to distinct binding pockets. We validated interactions between moesin and merlin FERM domains and full-length FAM83G, HIF1A, LATS1, NOP53, PAK6, RRBP1 and ZNF622 through pull-down experiments. Using biophysical binding assays, we determined affinities of, and uncovered allosteric interdependencies between, different binding partners, suggesting that the FERM domain acts as a switchable interaction hub. Using Rosetta FlexPepDock computational peptide docking protocols, we investigated the energy landscapes of identified interactions, which provide a detailed molecular understanding of the binding of the distinct binding motifs, as well as possible allosteric interconnections. This study demonstrates how experimental and computational approaches together can unravel a complex system of protein-peptide interactions that includes a family of proteins with multiple binding sites that interact with similar but distinct binding motifs.HighlightsWe screened the human disorderome for motif-containing partners of the FERM domainsWe expand the ERM and merlin interactomes of the ERMs and merlinWe identify four distinct motif classes that bind the ERM and merlin FERM domains: FYDF, xYxV, FY(D/E)L and LQE(I/L)In-vitro and in-silico data suggest that the FYDF motif binds to the F3a site and that xYxV motif binds to the F3b siteIn-silico modelling sheds light on the underlying conformational changes responsible for ligand interdependenciesAbstract Figure

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“…IQGAP1 is linked to the cortex via S100P and the plasma membrane- and actin-binding protein ezrin 48, 49 ( Figure 1). S100 proteins bind to Ca 2+ and the interaction between S100P and IQGAP1 is strictly Ca 2+ -dependent 48 .…”

Section: Coordination Of Cytoskeletal Cortical Interactions and Secrementioning

confidence: 99%

“…S100 proteins bind to Ca 2+ and the interaction between S100P and IQGAP1 is strictly Ca 2+ -dependent 48 . Also, ezrin has been shown to bind to Ca 2+ -bound S100P and IQGAP1, but since ezrin and IQGAP1 do interact in the absence of Ca 2+ , this interaction appeared to be S100P-independent 49 . Both S100P and ezrin co-localize with IQGAP1 in the cortical cell regions, and ezrin depletion reduced the cortical localization of IQGAP1 48, 49 .…”

Section: Coordination Of Cytoskeletal Cortical Interactions and Secrementioning

confidence: 99%

“…Also, ezrin has been shown to bind to Ca 2+ -bound S100P and IQGAP1, but since ezrin and IQGAP1 do interact in the absence of Ca 2+ , this interaction appeared to be S100P-independent 49 . Both S100P and ezrin co-localize with IQGAP1 in the cortical cell regions, and ezrin depletion reduced the cortical localization of IQGAP1 48, 49 . Interestingly, ezrin also interacts with the APC-binding protein Dlg1 50 , but it is not known whether APC, Dlg1, IQGAP1, and ezrin can function in the same complex.…”

Section: Coordination Of Cytoskeletal Cortical Interactions and Secrementioning

confidence: 99%

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Linking cortical microtubule attachment and exocytosis

Noordstra

Akhmanova

2017

F1000Res

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Exocytosis is a fundamental cellular process whereby secreted molecules are packaged into vesicles that move along cytoskeletal filaments and fuse with the plasma membrane. To function optimally, cells are strongly dependent on precisely controlled delivery of exocytotic cargo. In mammalian cells, microtubules serve as major tracks for vesicle transport by motor proteins, and thus microtubule organization is important for targeted delivery of secretory carriers. Over the years, multiple microtubule-associated and cortical proteins have been discovered that facilitate the interaction between the microtubule plus ends and the cell cortex. In this review, we focus on mammalian protein complexes that have been shown to participate in both cortical microtubule capture and exocytosis, thereby regulating the spatial organization of secretion. These complexes include microtubule plus-end tracking proteins, scaffolding factors, actin-binding proteins, and components of vesicle docking machinery, which together allow efficient coordination of cargo transport and release.

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Ezrin interacts with the scaffold protein IQGAP1 and affects its cortical localization

Cited by 10 publications

References 25 publications

Identification of IQGAP1 as a SLC26A4 (Pendrin)-Binding Protein in the Kidney

Identification of IQGAP1 as a SLC26A4 (Pendrin)-Binding Protein in the Kidney

Short linear motif based interactions and dynamics of the ezrin, radixin, moesin and merlin FERM domains

Linking cortical microtubule attachment and exocytosis

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