scholarly journals Redox protein Memo1 coordinates FGF23-driven signaling and small Rho-GTPases in the mouse kidney

2020 ◽  
Author(s):  
Katalin Bartos ◽  
Suresh Krishna Ramakrishnan ◽  
Sophie Braga-Lagache ◽  
Barbara Hänzi ◽  
Fanny Durussel ◽  
...  
Keyword(s):  
Rho Gtpases ◽  
Rho Gtpase ◽  
Premature Aging ◽  
Loss Of Function ◽  
Transcriptional Responses ◽  
Redox Protein ◽  
Rtk Signaling ◽  
Erk Phosphorylation ◽  
Small Rho Gtpases ◽  
Biochemical Analyses

AbstractMemo promotes receptor tyrosine kinase (RTK) signaling by unknown mechanisms. Memo1 deletion in mice causes premature aging and unbalanced metabolism partially resembling Fgf23 and Klotho loss-of-function animals. Here, we report a role for Memo’s redox function in FGF23-driven RTK signaling in the kidney. Postnatally Memo-deficient (cKO) and floxed controls were treated with FGF23 or vehicle, followed by molecular and biochemical analyses. Findings were validated using cell culture and recombinant proteins. Memo cKO mice showed impaired renal ERK phosphorylation and transcriptional responses to FGF23. Redox proteomics revealed excessive thiols of Rho-GDP dissociation inhibitor 1 (Rho-GDI1). Renal RhoA abundance and activity were increased in Memo cKO. Immunoprecipitation analysis showed an association between Memo and Rho-GDI1. We confirmed an interaction between the two proteins, with Memo-dependent irreversible oxidation at Rho-GDI1 Cys79 in cell-free conditions. Collectively, our findings reveal that redox protein Memo promotes renal FGF23 signaling together with oxidative modulation of the Rho-GTPase network.

Loss of Rho-GDIα sensitizes podocytes to lipopolysaccharide-mediated injury

2015 ◽  
Vol 308 (11) ◽  
pp. F1207-F1216 ◽  
Author(s):  
Richard Robins ◽  
Cindy Baldwin ◽  
Lamine Aoudjit ◽  
Indra R. Gupta ◽  
Tomoko Takano
Keyword(s):  
Nephrotic Syndrome ◽  
Rho Gtpases ◽  
Rho Gtpase ◽  
Cathepsin L ◽  
Loss Of Function ◽  
Filtration Barrier ◽  
Guanine Nucleotide Dissociation Inhibitor ◽  
Ubiquitin Proteasome ◽  
Proteasome Pathway ◽  
Lps Treatment

Nephrotic syndrome is a disease of glomerular permselectivity that can arise as a consequence of heritable or acquired changes to the integrity of the glomerular filtration barrier. We recently reported two siblings with heritable nephrotic syndrome caused by a loss of function mutation in the gene ARHGDIA, which encodes for Rho guanine nucleotide dissociation inhibitor-α (GDIα). GDIs are known to negatively regulate Rho-GTPase signaling. We hypothesized that loss of GDIα sensitizes podocytes to external injury via hyperactivation of Rho-GTPases and p38 MAPK. We examined the response of cultured podocytes with and without knockdown of GDIα to LPS injury by assessing the levels of phospho-p38 as well as the degree of synaptopodin loss. GDIα knockdown podocytes showed more pronounced and sustained p38 phosphorylation in response to LPS compared with control podocytes, and this was blunted significantly by the Rac1 inhibitor. In LPS-treated control podocytes, synaptopodin degradation occurred, and this was dependent on p38, the proteasome, and cathepsin L. In GDIα knockdown podocytes, the same events were triggered, but the levels of synaptopodin after LPS treatment were significantly lower than in control podocytes. These experiments reveal a common pathway by which heritable and environmental risk factors converge to injure podocytes, from Rac1 hyperactivation to p38 phosphorylation and synaptopodin degradation via the ubiquitin-proteasome pathway and cathepsin L.

scholarly journals The Exo70 Subunit of the Exocyst Is an Effector for Both Cdc42 and Rho3 Function in Polarized Exocytosis

2010 ◽  
Vol 21 (3) ◽  
pp. 430-442 ◽  
Author(s):  
Hao Wu ◽  
Courtney Turner ◽  
Jimmy Gardner ◽  
Brenda Temple ◽  
Patrick Brennwald
Keyword(s):  
Rho Gtpases ◽  
Rho Gtpase ◽  
Biochemical Properties ◽  
Loss Of Function ◽  
Gain Of Function ◽  
Present Evidence ◽  
Spatial Regulation ◽  
Exocyst Complex ◽  
Polarized Exocytosis

The Rho3 and Cdc42 members of the Rho GTPase family are important regulators of exocytosis in yeast. However, the precise mechanism by which they regulate this process is controversial. Here, we present evidence that the Exo70 component of the exocyst complex is a direct effector of both Rho3 and Cdc42. We identify gain-of-function mutants in EXO70 that potently suppress mutants in RHO3 and CDC42 defective for exocytic function. We show that Exo70 has the biochemical properties expected of a direct effector for both Rho3 and Cdc42. Surprisingly, we find that C-terminal prenylation of these GTPases both promotes the interaction and influences the sites of binding within Exo70. Finally, we demonstrate that the phenotypes associated with novel loss-of-function mutants in EXO70, are entirely consistent with Exo70 as an effector for both Rho3 and Cdc42 function in secretion. These data suggest that interaction with the Exo70 component of the exocyst is a key event in spatial regulation of exocytosis by Rho GTPases.

scholarly journals An RNAi screen unravels the complexities of Rho GTPase networks in skin morphogenesis

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Melanie Laurin ◽  
Nicholas C Gomez ◽  
John Levorse ◽  
Ataman Sendoel ◽  
Megan Sribour ◽  
...  
Keyword(s):  
Cell Polarity ◽  
Rho Gtpases ◽  
Planar Cell Polarity ◽  
Rho Gtpase ◽  
Gene Families ◽  
Tissue Imaging ◽  
Loss Of Function ◽  
Cytoskeletal Dynamics ◽  
Skin Morphogenesis ◽  
Gain Loss

During mammalian embryogenesis, extensive cellular remodeling is needed for tissue morphogenesis. As effectors of cytoskeletal dynamics, Rho GTPases and their regulators are likely involved, but their daunting complexity has hindered progress in dissecting their functions. We overcome this hurdle by employing high throughput in utero RNAi-mediated screening to identify key Rho regulators of skin morphogenesis. Our screen unveiled hitherto unrecognized roles for Rho-mediated cytoskeletal remodeling events that impact hair follicle specification, differentiation, downgrowth and planar cell polarity. Coupling our top hit with gain/loss-of-function genetics, interactome proteomics and tissue imaging, we show that RHOU, an atypical Rho, governs the cytoskeletal-junction dynamics that establish columnar shape and planar cell polarity in epidermal progenitors. Conversely, RHOU downregulation is required to remodel to a conical cellular shape that enables hair bud invagination and downgrowth. Our findings underscore the power of coupling screens with proteomics to unravel the physiological significance of complex gene families.

scholarly journals SRGAP1 Controls Small Rho GTPases To Regulate Podocyte Foot Process Maintenance

2021 ◽  
Vol 32 (3) ◽  
pp. 563-579
Author(s):  
Manuel Rogg ◽  
Jasmin I. Maier ◽  
Robert Dotzauer ◽  
Nadine Artelt ◽  
Oliver Kretz ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Rho Gtpases ◽  
Rho Gtpase ◽  
Conditional Knockout ◽  
Glomerular Diseases ◽  
Actin Networks ◽  
Small Rho Gtpases ◽  
Foot Process

BackgroundPrevious research demonstrated that small Rho GTPases, modulators of the actin cytoskeleton, are drivers of podocyte foot-process effacement in glomerular diseases, such as FSGS. However, a comprehensive understanding of the regulatory networks of small Rho GTPases in podocytes is lacking.MethodsWe conducted an analysis of podocyte transcriptome and proteome datasets for Rho GTPases; mapped in vivo, podocyte-specific Rho GTPase affinity networks; and examined conditional knockout mice and murine disease models targeting Srgap1. To evaluate podocyte foot-process morphology, we used super-resolution microscopy and electron microscopy; in situ proximity ligation assays were used to determine the subcellular localization of the small GTPase-activating protein SRGAP1. We performed functional analysis of CRISPR/Cas9-generated SRGAP1 knockout podocytes in two-dimensional and three-dimensional cultures and quantitative interaction proteomics.ResultsWe demonstrated SRGAP1 localization to podocyte foot processes in vivo and to cellular protrusions in vitro. Srgap1fl/fl*Six2Cre but not Srgap1fl/fl*hNPHS2Cre knockout mice developed an FSGS-like phenotype at adulthood. Podocyte-specific deletion of Srgap1 by hNPHS2Cre resulted in increased susceptibility to doxorubicin-induced nephropathy. Detailed analysis demonstrated significant effacement of podocyte foot processes. Furthermore, SRGAP1-knockout podocytes showed excessive protrusion formation and disinhibition of the small Rho GTPase machinery in vitro. Evaluation of a SRGAP1-dependent interactome revealed the involvement of SRGAP1 with protrusive and contractile actin networks. Analysis of glomerular biopsy specimens translated these findings toward human disease by displaying a pronounced redistribution of SRGAP1 in FSGS.ConclusionsSRGAP1, a podocyte-specific RhoGAP, controls podocyte foot-process architecture by limiting the activity of protrusive, branched actin networks. Therefore, elucidating the complex regulatory small Rho GTPase affinity network points to novel targets for potentially precise intervention in glomerular diseases.

scholarly journals Rho GTPase function in flies: insights from a developmental and organismal perspective

2004 ◽  
Vol 82 (6) ◽  
pp. 643-657 ◽  
Author(s):  
James E Johndrow ◽  
Craig R Magie ◽  
Susan M Parkhurst
Keyword(s):  
Rho Gtpases ◽  
Rho Gtpase ◽  
Fruit Fly ◽  
Genetic System ◽  
Small Gtpases ◽  
Dominant Negative ◽  
Loss Of Function ◽  
Cellular Behavior ◽  
Essential Components ◽  
Work Done

Morphogenesis is a key event in the development of a multicellular organism and is reliant on coordinated transcriptional and signal transduction events. To establish the segmented body plan that underlies much of metazoan development, individual cells and groups of cells must respond to exogenous signals with complex movements and shape changes. One class of proteins that plays a pivotal role in the interpretation of extracellular cues into cellular behavior is the Rho family of small GTPases. These molecular switches are essential components of a growing number of signaling pathways, many of which regulate actin cytoskeletal remodeling. Much of our understanding of Rho biology has come from work done in cell culture. More recently, the fruit fly Drosophila melanogaster has emerged as an excellent genetic system for the study of these proteins in a developmental and organismal context. Studies in flies have greatly enhanced our understanding of pathways involving Rho GTPases and their roles in development.Key words: Rho GTPases, Drosophila, development, effectors, loss-of-function mutation, dominant-negative, constitutively active.

scholarly journals The atypical Rho GTPase Rnd2 is critical for dentate granule neuron development and anxiety-like behavior during adult but not neonatal neurogenesis

2021 ◽  
Author(s):  
Thomas Kerloch ◽  
Fanny Farrugia ◽  
Lou Bouit ◽  
Marlène Maître ◽  
Geoffrey Terral ◽  
...  
Keyword(s):  
Adult Neurogenesis ◽  
Rho Gtpases ◽  
Neuronal Development ◽  
Rho Gtpase ◽  
Hippocampal Neurogenesis ◽  
Adult Hippocampal Neurogenesis ◽  
Granule Neurons ◽  
Loss Of Function ◽  
Neuron Development

AbstractDespite the central role of Rho GTPases in neuronal development, their functions in adult hippocampal neurogenesis remain poorly explored. Here, by using a retrovirus-based loss-of-function approach in vivo, we show that the atypical Rho GTPase Rnd2 is crucial for survival, positioning, somatodendritic morphogenesis, and functional maturation of adult-born dentate granule neurons. Interestingly, most of these functions are specific to granule neurons generated during adulthood since the deletion of Rnd2 in neonatally-born granule neurons only affects dendritogenesis. In addition, suppression of Rnd2 in adult-born dentate granule neurons increases anxiety-like behavior whereas its deletion in pups has no such effect, a finding supporting the adult neurogenesis hypothesis of anxiety disorders. Thus, our results are in line with the view that adult neurogenesis is not a simple continuation of earlier processes from development, and establish a causal relationship between Rnd2 expression and anxiety.

scholarly journals Selectivity Determinants of RHO GTPase Binding to IQGAPs

2021 ◽  
Vol 22 (22) ◽  
pp. 12596
Author(s):  
Niloufar Mosaddeghzadeh ◽  
Kazem Nouri ◽  
Oliver H. F. Krumbach ◽  
Ehsan Amin ◽  
Radovan Dvorsky ◽  
...  
Keyword(s):  
Rho Gtpases ◽  
Rho Gtpase ◽  
Decisive Role ◽  
Effector Proteins ◽  
Iq Motif ◽  
Cellular Processes ◽  
Wide Range ◽  
Switch Regions ◽  
Protein Components ◽  
Biochemical Analyses

IQ motif-containing GTPase-activating proteins (IQGAPs) modulate a wide range of cellular processes by acting as scaffolds and driving protein components into distinct signaling networks. Their functional states have been proposed to be controlled by members of the RHO family of GTPases, among other regulators. In this study, we show that IQGAP1 and IQGAP2 can associate with CDC42 and RAC1-like proteins but not with RIF, RHOD, or RHO-like proteins, including RHOA. This seems to be based on the distribution of charged surface residues, which varies significantly among RHO GTPases despite their high sequence homology. Although effector proteins bind first to the highly flexible switch regions of RHO GTPases, additional contacts outside are required for effector activation. Sequence alignment and structural, mutational, and competitive biochemical analyses revealed that RHO GTPases possess paralog-specific residues outside the two highly conserved switch regions that essentially determine the selectivity of RHO GTPase binding to IQGAPs. Amino acid substitution of these specific residues in RHOA to the corresponding residues in RAC1 resulted in RHOA association with IQGAP1. Thus, electrostatics most likely plays a decisive role in these interactions.

The role of RhoJ in endothelial cell biology and angiogenesis

2011 ◽  
Vol 39 (6) ◽  
pp. 1606-1611 ◽  
Author(s):  
Katarzyna Leszczynska ◽  
Sukhbir Kaur ◽  
Eleanor Wilson ◽  
Roy Bicknell ◽  
Victoria L. Heath
Keyword(s):  
Focal Adhesion ◽  
Rho Gtpases ◽  
Cell Biology ◽  
Rho Gtpase ◽  
Vascular Endothelial ◽  
Small Rho Gtpases ◽  
Endothelial Growth Factor

RhoJ is an endothelially expressed member of the Cdc42 (cell division cycle 42) subfamily of small Rho GTPases. It is expressed in both the developing mammalian vasculature and the vascular beds of a number of adult tissues, with its expression regulated by the endothelial transcription factor ERG (ETS-related gene). RhoJ has been shown to regulate endothelial motility, tubulogenesis and lumen formation in vitro, and modulates the vascularization of Matrigel plugs in vivo. Both vascular endothelial growth factor and semaphorin 3E have been found to affect its activation. RhoJ has been shown to be a focal-adhesion-localized Rho GTPase which can modulate focal adhesion number, actomyosin contractility and activity of Cdc42 and Rac1. The present review discusses the biology of RhoJ with a focus on recent reports of its role in endothelial cells and angiogenesis.

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