Rab35 Mediates Transport of Cdc42 and Rac1 to the Plasma Membrane during Phagocytosis

ABSTRACT Phagocytosis of invading microbes requires dynamic rearrangement of the plasma membrane and its associated cytoskeletal actin network. The polarization of Cdc42 and Rac1 Rho GTPases to the site of plasma membrane protrusion is responsible for the remodeling of actin structures. However, the mechanism of Rho GTPase recruitment to these sites and the identities of accessory molecules involved in this process are not well understood. In this study, we uncovered several new components involved in innate immunity in Drosophila melanogaster. Our data demonstrate that Rab35 is a regulator of vesicle transport required specifically for phagocytosis. Moreover, recruitment of Cdc42 and Rac1 to the sites of filopodium and lamellipodium formation is Rab35 dependent and occurs by way of microtubule tracks. These results implicate Rab35 as the immune cell-specific regulator of vesicle transport within the actin-remodeling complex.

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The TRE17 Oncogene Encodes a Component of a Novel Effector Pathway for Rho GTPases Cdc42 and Rac1 and Stimulates Actin Remodeling

Molecular and Cellular Biology ◽

10.1128/mcb.23.6.2151-2161.2003 ◽

2003 ◽

Vol 23 (6) ◽

pp. 2151-2161 ◽

Cited By ~ 36

Author(s):

Jeffrey M. Masuda-Robens ◽

Sara N. Kutney ◽

Hongwei Qi ◽

Margaret M. Chou

Keyword(s):

Plasma Membrane ◽

Rho Gtpases ◽

Actin Polymerization ◽

Dependent Manner ◽

Cortical Actin ◽

Actin Remodeling ◽

Membrane Recruitment ◽

Truncation Mutant ◽

Effector Pathway

ABSTRACT The Rho family GTPases Cdc42 and Rac1 play fundamental roles in transformation and actin remodeling. Here, we demonstrate that the TRE17 oncogene encodes a component of a novel effector pathway for these GTPases. TRE17 coprecipitated specifically with the active forms of Cdc42 and Rac1 in vivo. Furthermore, the subcellular localization of TRE17 was dramatically regulated by these GTPases and mitogens. Under serum-starved conditions, TRE17 localized predominantly to filamentous structures within the cell. Epidermal growth factor (EGF) induced relocalization of TRE17 to the plasma membrane in a Cdc42-/Rac1-dependent manner. Coexpression of activated alleles of Cdc42 or Rac1 also caused complete redistribution of TRE17 to the plasma membrane, where it partially colocalized with the GTPases in filopodia and ruffles, respectively. Membrane recruitment of TRE17 by EGF or the GTPases was dependent on actin polymerization. Finally, we found that a C-terminal truncation mutant of TRE17 induced the accumulation of cortical actin, mimicking the effects of activated Cdc42. Together, these results identify TRE17 as part of a novel effector complex for Cdc42 and Rac1, potentially contributing to their effects on actin remodeling. The present study provides insights into the regulation and cellular function of this previously uncharacterized oncogene.

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Functional and Therapeutic Relevance of Rho GTPases in Innate Immune Cell Migration and Function during Inflammation: An In Silico Perspective

Mediators of Inflammation ◽

10.1155/2021/6655412 ◽

2021 ◽

Vol 2021 ◽

pp. 1-10

Author(s):

Pankaj Dipankar ◽

Puneet Kumar ◽

Shiba Prasad Dash ◽

Pranita P. Sarangi

Keyword(s):

Cell Migration ◽

Immune Cells ◽

Rho Gtpases ◽

Inflammatory Cell ◽

Immune Cell ◽

Rho Gtpase ◽

Innate Immune ◽

Molecular Organization ◽

Exchange Reactions ◽

And Function

Systematic regulation of leukocyte migration to the site of infection is a vital step during immunological responses. Improper migration and localization of immune cells could be associated with disease pathology as seen in systemic inflammation. Rho GTPases act as molecular switches during inflammatory cell migration by cycling between Rho-GDP (inactive) to Rho-GTP (active) forms and play an essential role in the precise regulation of actin cytoskeletal dynamics as well as other immunological functions of leukocytes. Available reports suggest that the dysregulation of Rho GTPase signaling is associated with various inflammatory diseases ranging from mild to life-threatening conditions. Therefore, it is crucial to understand the step-by-step activation and inactivation of GTPases and the functioning of different Guanine Nucleotide Exchange Factors (GEFs) and GTPase-Activating Proteins (GAPs) that regulate the conversion of GDP to GTP and GTP to GDP exchange reactions, respectively. Here, we describe the molecular organization and activation of various domains of crucial elements associated with the activation of Rho GTPases using solved PDB structures. We will also present the latest evidence available on the relevance of Rho GTPases in the migration and function of innate immune cells during inflammation. This knowledge will help scientists design promising drug candidates against the Rho-GTPase-centric regulatory molecules regulating inflammatory cell migration.

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Plasma membrane nano-organization specifies phosphoinositide effects on Rho-GTPases and actin dynamics in tobacco pollen tubes

10.1101/2020.08.12.248419 ◽

2020 ◽

Cited By ~ 1

Author(s):

Marta Fratini ◽

Praveen Krishnamoorthy ◽

Irene Stenzel ◽

Mara Riechmann ◽

Kirsten Bacia ◽

...

Keyword(s):

Plasma Membrane ◽

Membrane Trafficking ◽

Rho Gtpases ◽

Rho Gtpase ◽

Pollen Tubes ◽

Actin Dynamics ◽

Tobacco Pollen ◽

Dual Distribution ◽

Cytoskeletal Dynamics

AbstractPollen tube growth requires coordination of cytoskeletal dynamics and apical secretion. The regulatory phospholipid, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) is enriched in the subapical plasma membrane of pollen tubes and can influence both actin dynamics and secretion. How alternative PtdIns(4,5)P2-effects are specified is unclear. Spinning disc microscopy (SD) reveals dual distribution of a fluorescent PtdIns(4,5)P2-reporter in dynamic plasma membrane nanodomains vs. apparent diffuse membrane labelling, consistent with spatially distinct coexisting pools of PtdIns(4,5)P2. Several PI4P 5-kinases (PIP5Ks) can generate PtdIns(4,5)P2 in pollen tubes. Despite localizing to one membrane region, AtPIP5K2 and NtPIP5K6 display distinctive overexpression effects on cell morphologies, respectively related to altered actin dynamics or membrane trafficking. When analyzed by SD, AtPIP5K2-EYFP associated with nanodomains, whereas NtPIP5K6-EYFP localized diffusely. Chimeric AtPIP5K2 and NtPIP5K6 variants with reciprocally swapped membrane-associating domains evoked reciprocally shifted effects on cell morphology upon overexpression. Overall, PI4P 5-kinase variants targeted to nanodomains stabilized actin, suggesting a specific function of PtdIns(4,5)P2-nanodomains. A distinct role of nanodomain-associated AtPIP5K2 in actin regulation is further supported by proximity to and interaction with the Rho-GTPase NtRac5, and by functional interplay with elements of ROP-signalling. Plasma membrane nano-organization may thus aid the specification of PtdIns(4,5)P2-functions to coordinate cytoskeletal dynamics and secretion in pollen tubes.

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Actin dynamics in cell migration

Essays in Biochemistry ◽

10.1042/ebc20190015 ◽

2019 ◽

Vol 63 (5) ◽

pp. 483-495 ◽

Cited By ~ 22

Author(s):

Matthias Schaks ◽

Grégory Giannone ◽

Klemens Rottner

Keyword(s):

Plasma Membrane ◽

Cell Migration ◽

Actin Filament ◽

Rho Gtpases ◽

Rho Gtpase ◽

Actin Dynamics ◽

Regulatory Complex ◽

Membrane Protrusions ◽

Filament Networks ◽

Contractile Forces

Abstract Cell migration is an essential process, both in unicellular organisms such as amoeba and as individual or collective motility in highly developed multicellular organisms like mammals. It is controlled by a variety of activities combining protrusive and contractile forces, normally generated by actin filaments. Here, we summarize actin filament assembly and turnover processes, and how respective biochemical activities translate into different protrusion types engaged in migration. These actin-based plasma membrane protrusions include actin-related protein 2/3 complex-dependent structures such as lamellipodia and membrane ruffles, filopodia as well as plasma membrane blebs. We also address observed antagonisms between these protrusion types, and propose a model – also inspired by previous literature – in which a complex balance between specific Rho GTPase signaling pathways dictates the protrusion mechanism employed by cells. Furthermore, we revisit published work regarding the fascinating antagonism between Rac and Rho GTPases, and how this intricate signaling network can define cell behavior and modes of migration. Finally, we discuss how the assembly of actin filament networks can feed back onto their regulators, as exemplified for the lamellipodial factor WAVE regulatory complex, tightly controlling accumulation of this complex at specific subcellular locations as well as its turnover.

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The Cytotoxic Necrotizing Factors (CNFs)—A Family of Rho GTPase-Activating Bacterial Exotoxins

Toxins ◽

10.3390/toxins13120901 ◽

2021 ◽

Vol 13 (12) ◽

pp. 901

Author(s):

Paweena Chaoprasid ◽

Petra Dersch

Keyword(s):

Rho Gtpases ◽

Pathogenic Bacteria ◽

Immune Cell ◽

Rho Gtpase ◽

Current Knowledge ◽

Single Chain ◽

Tissue Integrity ◽

Cell Tissue ◽

Functional Consequences ◽

Cell Processes

The cytotoxic necrotizing factors (CNFs) are a family of Rho GTPase-activating single-chain exotoxins that are produced by several Gram-negative pathogenic bacteria. Due to the pleiotropic activities of the targeted Rho GTPases, the CNFs trigger multiple signaling pathways and host cell processes with diverse functional consequences. They influence cytokinesis, tissue integrity, cell barriers, and cell death, as well as the induction of inflammatory and immune cell responses. This has an enormous influence on host–pathogen interactions and the severity of the infection. The present review provides a comprehensive insight into our current knowledge of the modular structure, cell entry mechanisms, and the mode of action of this class of toxins, and describes their influence on the cell, tissue/organ, and systems levels. In addition to their toxic functions, possibilities for their use as drug delivery tool and for therapeutic applications against important illnesses, including nervous system diseases and cancer, have also been identified and are discussed.

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Regulation of Rho GTPases by RhoGDIs in Human Cancers

Cells ◽

10.3390/cells8091037 ◽

2019 ◽

Vol 8 (9) ◽

pp. 1037 ◽

Cited By ~ 6

Author(s):

Cho ◽

Kim ◽

Baek ◽

Kim ◽

Lee

Keyword(s):

Cell Migration ◽

Cancer Progression ◽

Anticancer Agents ◽

Rho Gtpases ◽

Rho Gtpase ◽

Regulatory Mechanisms ◽

Malignant Phenotype ◽

Cellular Processes ◽

Human Cancers

Rho GDP dissociation inhibitors (RhoGDIs) play important roles in various cellular processes, including cell migration, adhesion, and proliferation, by regulating the functions of the Rho GTPase family. Dissociation of Rho GTPases from RhoGDIs is necessary for their spatiotemporal activation and is dynamically regulated by several mechanisms, such as phosphorylation, sumoylation, and protein interaction. The expression of RhoGDIs has changed in many human cancers and become associated with the malignant phenotype, including migration, invasion, metastasis, and resistance to anticancer agents. Here, we review how RhoGDIs control the function of Rho GTPases by regulating their spatiotemporal activity and describe the regulatory mechanisms of the dissociation of Rho GTPases from RhoGDIs. We also discuss the role of RhoGDIs in cancer progression and their potential uses for therapeutic intervention.

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Injury-Induced Innate Immune Response During Segment Regeneration of the Earthworm, Eisenia andrei

International Journal of Molecular Sciences ◽

10.3390/ijms22052363 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2363

Author(s):

Kornélia Bodó ◽

Zoltán Kellermayer ◽

Zoltán László ◽

Ákos Boros ◽

Bohdana Kokhanyuk ◽

...

Keyword(s):

Immune Response ◽

Innate Immunity ◽

Immune Cell ◽

Scavenger Receptor ◽

Cell Renewal ◽

Body Parts ◽

Eisenia Andrei ◽

Blastema Formation ◽

Close Proximity ◽

Renewal Period

Regeneration of body parts and their interaction with the immune response is a poorly understood aspect of earthworm biology. Consequently, we aimed to study the mechanisms of innate immunity during regeneration in Eisenia andrei earthworms. In the course of anterior and posterior regeneration, we documented the kinetical aspects of segment restoration by histochemistry. Cell proliferation peaked at two weeks and remitted by four weeks in regenerating earthworms. Apoptotic cells were present throughout the cell renewal period. Distinct immune cell (e.g., coelomocyte) subsets were accumulated in the newly-formed blastema in the close proximity of the apoptotic area. Regenerating earthworms have decreased pattern recognition receptors (PRRs) (e.g., TLR, except for scavenger receptor) and antimicrobial peptides (AMPs) (e.g., lysenin) mRNA patterns compared to intact earthworms. In contrast, at the protein level, mirroring regulation of lysenins became evident. Experimental coelomocyte depletion caused significantly impaired cell divisions and blastema formation during anterior and posterior regeneration. These obtained novel data allow us to gain insight into the intricate interactions of regeneration and invertebrate innate immunity.

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PAK and other Rho-associated kinases – effectors with surprisingly diverse mechanisms of regulation

Biochemical Journal ◽

10.1042/bj20041638 ◽

2005 ◽

Vol 386 (2) ◽

pp. 201-214 ◽

Cited By ~ 190

Author(s):

Zhou-shen ZHAO ◽

Ed MANSER

Keyword(s):

Rho Gtpases ◽

Cell Biology ◽

Rho Gtpase ◽

Rho Kinase ◽

Molecular Switches ◽

Eukaryotic Cell ◽

Effector Proteins ◽

Downstream Effector ◽

P21 Activated Kinase ◽

Do So

The Rho GTPases are a family of molecular switches that are critical regulators of signal transduction pathways in eukaryotic cells. They are known principally for their role in regulating the cytoskeleton, and do so by recruiting a variety of downstream effector proteins. Kinases form an important class of Rho effector, and part of the biological complexity brought about by switching on a single GTPase results from downstream phosphorylation cascades. Here we focus on our current understanding of the way in which different Rho-associated serine/threonine kinases, denoted PAK (p21-activated kinase), MLK (mixed-lineage kinase), ROK (Rho-kinase), MRCK (myotonin-related Cdc42-binding kinase), CRIK (citron kinase) and PKN (protein kinase novel), interact with and are regulated by their partner GTPases. All of these kinases have in common an ability to dimerize, and in most cases interact with a variety of other proteins that are important for their function. A diversity of known structures underpin the Rho GTPase–kinase interaction, but only in the case of PAK do we have a good molecular understanding of kinase regulation. The ability of Rho GTPases to co-ordinate spatial and temporal phosphorylation events explains in part their prominent role in eukaryotic cell biology.

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The BNIP-2 and Cdc42GAP Homology (BCH) Domain of p50RhoGAP/Cdc42GAP Sequesters RhoA from Inactivation by the Adjacent GTPase-activating Protein Domain

Molecular Biology of the Cell ◽

10.1091/mbc.e09-05-0408 ◽

2010 ◽

Vol 21 (18) ◽

pp. 3232-3246 ◽

Cited By ~ 13

Author(s):

Yi Ting Zhou ◽

Li Li Chew ◽

Sheng-cai Lin ◽

Boon Chuan Low

Keyword(s):

Rho Gtpases ◽

Rho Gtpase ◽

Intramolecular Interaction ◽

Complete Loss ◽

Protein Domain ◽

Binding Motif ◽

Gtpase Activating Protein ◽

New Paradigm ◽

Cell Rounding ◽

In Cis

The BNIP-2 and Cdc42GAP homology (BCH) domain is a novel regulator for Rho GTPases, but its impact on p50-Rho GTPase-activating protein (p50RhoGAP or Cdc42GAP) in cells remains elusive. Here we show that deletion of the BCH domain from p50RhoGAP enhanced its GAP activity and caused drastic cell rounding. Introducing constitutively active RhoA or inactivating GAP domain blocked such effect, whereas replacing the BCH domain with endosome-targeting SNX3 excluded requirement of endosomal localization in regulating the GAP activity. Substitution with homologous BCH domain from Schizosaccharomyces pombe, which does not bind mammalian RhoA, also led to complete loss of suppression. Interestingly, the p50RhoGAP BCH domain only targeted RhoA, but not Cdc42 or Rac1, and it was unable to distinguish between GDP and the GTP-bound form of RhoA. Further mutagenesis revealed a RhoA-binding motif (residues 85-120), which when deleted, significantly reduced BCH inhibition on GAP-mediated cell rounding, whereas its full suppression also required an intramolecular interaction motif (residues 169-197). Therefore, BCH domain serves as a local modulator in cis to sequester RhoA from inactivation by the adjacent GAP domain, adding to a new paradigm for regulating p50RhoGAP signaling.

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Post-Translational Modification and Subcellular Compartmentalization: Emerging Concepts on the Regulation and Physiopathological Relevance of RhoGTPases

Cells ◽

10.3390/cells10081990 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1990

Author(s):

Inmaculada Navarro-Lérida ◽

Miguel Sánchez-Álvarez ◽

Miguel Ángel del Pozo

Keyword(s):

Rho Gtpases ◽

Rho Gtpase ◽

Protein Complexes ◽

Specific Protein ◽

Cell Polarization ◽

Small Gtpase ◽

Post Translational Modification ◽

Functional Regulation ◽

Subcellular Compartmentalization ◽

Cellular Compartments

Cells and tissues are continuously exposed to both chemical and physical stimuli and dynamically adapt and respond to this variety of external cues to ensure cellular homeostasis, regulated development and tissue-specific differentiation. Alterations of these pathways promote disease progression—a prominent example being cancer. Rho GTPases are key regulators of the remodeling of cytoskeleton and cell membranes and their coordination and integration with different biological processes, including cell polarization and motility, as well as other signaling networks such as growth signaling and proliferation. Apart from the control of GTP–GDP cycling, Rho GTPase activity is spatially and temporally regulated by post-translation modifications (PTMs) and their assembly onto specific protein complexes, which determine their controlled activity at distinct cellular compartments. Although Rho GTPases were traditionally conceived as targeted from the cytosol to the plasma membrane to exert their activity, recent research demonstrates that active pools of different Rho GTPases also localize to endomembranes and the nucleus. In this review, we discuss how PTM-driven modulation of Rho GTPases provides a versatile mechanism for their compartmentalization and functional regulation. Understanding how the subcellular sorting of active small GTPase pools occurs and what its functional significance is could reveal novel therapeutic opportunities.

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