scholarly journals Co-ordinated Ras and Rac activity shapes macropinocytic cups and enables phagocytosis of geometrically diverse bacteria

2019 ◽  
Author(s):  
Catherine M. Buckley ◽  
Henderikus Pots ◽  
Aurelie Gueho ◽  
Ben A. Phillips ◽  
Bernd Gilsbach ◽  
...  
Keyword(s):  
Dictyostelium Discoideum ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Extracellular Material ◽  
Multidomain Protein ◽  
Complex Shapes ◽  
Rho Family ◽  
Phagocytic Cups ◽  
Interior Domain ◽  
Professional Phagocyte

AbstractEngulfment of extracellular material by phagocytosis or macropinocytosis depends on the ability of cells to generate specialised cup shaped protrusions. To effectively capture and internalise their targets, these cups are organised into a ring or ruffle of actin-driven protrusion encircling a non-protrusive interior domain. These functional domains depend on the combined activities of multiple Ras and Rho family small GTPases, but how their activities are integrated and differentially regulated over space and time is unknown. Here, we show that the amoeba Dictyostelium discoideum coordinates Ras and Rac activity using the multidomain protein RGBARG (RCC1, RhoGEF, BAR and RasGAP-containing protein). We find RGBARG uses a tripartite mechanism of Ras, Rac and phospholipid interactions to localise at the protruding edge and interface with the interior of both macropinocytic and phagocytic cups. There, RGBARG shapes the protrusion by driving Rac activation at the rim whilst suppressing expansion of the active Ras interior domain. Consequently, cells lacking RGBARG form enlarged, flat interior domains unable to generate large macropinosomes. During phagocytosis, we find that disruption of RGBARG causes a geometry-specific defect in engulfing rod-shaped bacteria and ellipsoidal beads. This demonstrates the importance of co-ordinating small GTPase activities during engulfment of more complex shapes and thus the full physiological range of microbes, and how this is achieved in a model professional phagocyte.

scholarly journals PseudoGTPase domains in p190RhoGAP proteins: a mini-review

2018 ◽  
Vol 46 (6) ◽  
pp. 1713-1720 ◽  
Author(s):  
Amy L. Stiegler ◽  
Titus J. Boggon
Keyword(s):  
Family Members ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Signaling Proteins ◽  
New Family ◽  
Gtpase Activating Proteins ◽  
Rho Family ◽  
Biochemical Analyses ◽  
Catalytic Cleft

Pseudoenzymes generally lack detectable catalytic activity despite adopting the overall protein fold of their catalytically competent counterparts, indeed ‘pseudo’ family members seem to be incorporated in all enzyme classes. The small GTPase enzymes are important signaling proteins, and recent studies have identified many new family members with noncanonical residues within the catalytic cleft, termed pseudoGTPases. To illustrate recent discoveries in the field, we use the p190RhoGAP proteins as an example. p190RhoGAP proteins (ARHGAP5 and ARHGAP35) are the most abundant GTPase activating proteins for the Rho family of small GTPases. These are key regulators of Rho signaling in processes such as cell migration, adhesion and cytokinesis. Structural biology has complemented and guided biochemical analyses for these proteins and has allowed discovery of two cryptic pseudoGTPase domains, and the re-classification of a third, previously identified, GTPase-fold domain as a pseudoGTPase. The three domains within p190RhoGAP proteins illustrate the diversity of this rapidly expanding pseudoGTPase group.

scholarly journals Identification and characterization of a new isoform of small GTPase RhoE

2020 ◽  
Vol 3 (1) ◽  
Author(s):  
Yuan Dai ◽  
Weijia Luo ◽  
Xiaojing Yue ◽  
Wencai Ma ◽  
Jing Wang ◽  
...  
Keyword(s):  
Rho Gtpase ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Biological Functions ◽  
Coding Region ◽  
Alternative Translation ◽  
Rho Family ◽  
Binding Capability ◽  
Identification And Characterization

Abstract The Rho family of GTPases consists of 20 members including RhoE. Here, we discover the existence of a short isoform of RhoE designated as RhoEα, the first Rho GTPase isoform generated from alternative translation. Translation of this new isoform is initiated from an alternative start site downstream of and in-frame with the coding region of the canonical RhoE. RhoEα exhibits a similar subcellular distribution while its protein stability is higher than RhoE. RhoEα contains binding capability to RhoE effectors ROCK1, p190RhoGAP and Syx. The distinct transcriptomes of cells with the expression of RhoE and RhoEα, respectively, are demonstrated. The data propose distinctive and overlapping biological functions of RhoEα compared to RhoE. In conclusion, this study reveals a new Rho GTPase isoform generated from alternative translation. The discovery provides a new scope of understanding the versatile functions of small GTPases and underlines the complexity and diverse roles of small GTPases.

scholarly journals Isoprenylcysteine Carboxy Methylation Is Essential for Development in Dictyostelium discoideum

2007 ◽  
Vol 18 (10) ◽  
pp. 4106-4118 ◽  
Author(s):  
Ying Chen ◽  
Kyle J. McQuade ◽  
Xiao-Juan Guan ◽  
Peter A. Thomason ◽  
Michael S. Wert ◽  
...  
Keyword(s):  
Dictyostelium Discoideum ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Intercellular Signaling ◽  
Wild Type ◽  
Protein Subunit ◽  
Encoding Gene ◽  
Carboxy Terminal ◽  
Mutant Cells ◽  
Do So

Members of the Ras superfamily of small GTPases and the heterotrimeric G protein γ subunit are methylated on their carboxy-terminal cysteine residues by isoprenylcysteine methyltransferase. In Dictyostelium discoideum, small GTPase methylation occurs seconds after stimulation of starving cells by cAMP and returns quickly to basal levels, suggesting an important role in cAMP-dependent signaling. Deleting the isoprenylcysteine methyltransferase-encoding gene causes dramatic defects. Starving mutant cells do not propagate cAMP waves in a sustained manner, and they do not aggregate. Motility is rescued when cells are pulsed with exogenous cAMP, or coplated with wild-type cells, but the rescued cells exhibit altered polarity. cAMP-pulsed methyltransferase-deficient cells that have aggregated fail to differentiate, but mutant cells plated in a wild-type background are able to do so. Localization of and signaling by RasG is altered in the mutant. Localization of the heterotrimeric Gγ protein subunit was normal, but signaling was altered in mutant cells. These data indicate that isoprenylcysteine methylation is required for intercellular signaling and development in Dictyostelium.

scholarly journals Small GTPase RhoG Is a Key Regulator for Neurite Outgrowth in PC12 Cells

2000 ◽  
Vol 20 (19) ◽  
pp. 7378-7387 ◽  
Author(s):  
Hironori Katoh ◽  
Hidekazu Yasui ◽  
Yoshiaki Yamaguchi ◽  
Junko Aoki ◽  
Hirotada Fujita ◽  
...  
Keyword(s):  
Neurite Outgrowth ◽  
Pc12 Cells ◽  
Morphological Changes ◽  
Cell Types ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Dominant Negative ◽  
Wild Type ◽  
Rho Family ◽  
Constitutively Active

ABSTRACT The Rho family of small GTPases has been implicated in cytoskeletal reorganization and subsequent morphological changes in various cell types. Among them, Rac and Cdc42 have been shown to be involved in neurite outgrowth in neuronal cells. In this study, we examined the role of RhoG, another member of Rho family GTPases, in nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells. Expression of wild-type RhoG in PC12 cells induced neurite outgrowth in the absence of NGF, and the morphology of wild-type RhoG-expressing cells was similar to that of NGF-differentiated cells. Constitutively active RhoG-transfected cells extended short neurites but developed large lamellipodial or filopodial structures at the tips of neurites. RhoG-induced neurite outgrowth was inhibited by coexpression with dominant-negative Rac1 or Cdc42. In addition, expression of constitutively active RhoG elevated endogenous Rac1 and Cdc42 activities. We also found that the NGF-induced neurite outgrowth was enhanced by expression of wild-type RhoG whereas expression of dominant-negative RhoG suppressed the neurite outgrowth. Furthermore, constitutively active Ras-induced neurite outgrowth was also suppressed by dominant-negative RhoG. Taken together, these results suggest that RhoG is a key regulator in NGF-induced neurite outgrowth, acting downstream of Ras and upstream of Rac1 and Cdc42 in PC12 cells.

scholarly journals The discovery and maturation of peptide biologics targeting the small G-protein Cdc42: A bioblockade for Ras-driven signaling

2020 ◽  
Vol 295 (9) ◽  
pp. 2866-2884 ◽  
Author(s):  
George J. N. Tetley ◽  
Natasha P. Murphy ◽  
Stephane Bonetto ◽  
Gabriela Ivanova-Berndt ◽  
Jefferson Revell ◽  
...  
Keyword(s):  
Cyclic Peptide ◽  
Inhibitory Effect ◽  
Small Gtpases ◽  
Affinity Maturation ◽  
Small Gtpase ◽  
Ras Signaling ◽  
Cell Interior ◽  
Small G Protein ◽  
A Cell ◽  
Rho Family

Aberrant Ras signaling drives 30% of cancers, and inhibition of the Rho family small GTPase signaling has been shown to combat Ras-driven cancers. Here, we present the discovery of a 16-mer cyclic peptide that binds to Cdc42 with nanomolar affinity. Affinity maturation of this sequence has produced a panel of derived candidates with increased affinity and modulated specificity for other closely-related small GTPases. The structure of the tightest binding peptide was solved by NMR, and its binding site on Cdc42 was determined. Addition of a cell-penetrating sequence allowed the peptides to access the cell interior and engage with their target(s), modulating signaling pathways. In Ras-driven cancer cell models, the peptides have an inhibitory effect on proliferation and show suppression of both invasion and motility. As such, they represent promising candidates for Rho-family small GTPase inhibitors and therapeutics targeting Ras-driven cancers. Our data add to the growing literature demonstrating that peptides are establishing their place in the biologics arm of drug discovery.

scholarly journals S-Palmitoylation-Dependent Regulation of Cardiomyocyte Rac1 Signaling Activity and Cardiac Hypertrophy

2021 ◽  
Author(s):  
Matthew J Brody ◽  
Tanya A. Baldwin ◽  
Arasakumar Subramani ◽  
Onur Kanisicak ◽  
Ronald J Vagnozzi ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Transgenic Mice ◽  
Cardiac Disease ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Transferase Activity ◽  
Related Enzyme ◽  
Rho Family ◽  
Novel Therapeutic Strategies

S-palmitoylation is a reversible lipid modification that regulates trafficking, localization, activity, and/or stability of protein substrates by serving as a fatty acid anchor to cell membranes. However, S-palmitoylation-dependent control of signal transduction in cardiomyocytes and its effects on cardiac physiology are not well understood. We performed an in vivo gain-of-function screen of zinc finger Asp-His-His-Cys (zDHHC) family S-acyl transferases that catalyze S-palmitoylation and identified the Golgi-localized enzyme zDHHC3 as a critical regulator of cardiac maladaptation. The closely-related enzyme, zDHHC7, also induced severe cardiomyopathy but this effect was not observed with overexpression of plasma membrane enzyme zDHHC5, endoplasmic reticulum enzyme zDHHC6, or Golgi enzyme zDHHC13. To identify effectors that may underlie zDHHC3-induced cardiomyopathy we performed quantitative site-specific S-acyl proteomics in zDHHC3-overexpressing cells that revealed the small GTPase Rac1 as a novel substrate. We generated cardiomyocyte-specific transgenic mice overexpressing zDHHC3, which develop severe cardiac disease. Cardiomyopathy and congestive heart failure in zDHHC3 transgenic mice are preceded by enhanced S-palmitoylation of Rac1 and induction of additional Rho family small GTPases including RhoA, Cdc42, and the Rho family-specific chaperone RhoGDI. In contrast, transgenic mice overexpressing an enzymatically-dead mutant of zDHHC3 do not exhibit this profound induction of RhoGTPase signaling or develop cardiac disease. Rac1 S-palmitoylation, plasma membrane localization, activity, and downstream hypertrophic signaling were substantially increased in zDHHC3 overexpressing hearts. Taken together, these data suggest inhibition of zDHHC3/7 S-acyl transferase activity at the cardiomyocyte Golgi or disruption of Rac1 S-palmitoylation as novel therapeutic strategies to treat cardiac disease or other diseases associated with enhanced RhoGTPase signaling.

scholarly journals Post-Translational Modification and Subcellular Distribution of Rac1: An Update

Cells ◽  
2018 ◽  
Vol 7 (12) ◽  
pp. 263 ◽  
Author(s):  
Abdalla Abdrabou ◽  
Zhixiang Wang
Keyword(s):  
Membrane Trafficking ◽  
Subcellular Distribution ◽  
Bound States ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Guanine Nucleotide ◽  
Post Translational Modification ◽  
Post Translational Modifications ◽  
Rho Family ◽  
And Function

Rac1 is a small GTPase that belongs to the Rho family. The Rho family of small GTPases is a subfamily of the Ras superfamily. The Rho family of GTPases mediate a plethora of cellular effects, including regulation of cytoarchitecture, cell size, cell adhesion, cell polarity, cell motility, proliferation, apoptosis/survival, and membrane trafficking. The cycling of Rac1 between the GTP (guanosine triphosphate)- and GDP (guanosine diphosphate)-bound states is essential for effective signal flow to elicit downstream biological functions. The cycle between inactive and active forms is controlled by three classes of regulatory proteins: Guanine nucleotide exchange factors (GEFs), GTPase-activating proteins (GAPs), and guanine-nucleotide-dissociation inhibitors (GDIs). Other modifications include RNA splicing and microRNAs; various post-translational modifications have also been shown to regulate the activity and function of Rac1. The reported post-translational modifications include lipidation, ubiquitination, phosphorylation, and adenylylation, which have all been shown to play important roles in the regulation of Rac1 and other Rho GTPases. Moreover, the Rac1 activity and function are regulated by its subcellular distribution and translocation. This review focused on the most recent progress in Rac1 research, especially in the area of post-translational modification and subcellular distribution and translocation.

TBC proteins: GAPs for mammalian small GTPase Rab?

2011 ◽  
Vol 31 (3) ◽  
pp. 159-168 ◽  
Author(s):  
Mitsunori Fukuda
Keyword(s):  
Insulin Resistance ◽  
Small Gtpases ◽  
Structure And Function ◽  
Small Gtpase ◽  
Cell Cycle Regulators ◽  
Gtpase Activating Protein ◽  
Domain Family ◽  
Protein Motif ◽  
Conserved Domain ◽  
And Function

The TBC (Tre-2/Bub2/Cdc16) domain was originally identified as a conserved domain among the tre-2 oncogene product and the yeast cell cycle regulators Bub2 and Cdc16, and it is now widely recognized as a conserved protein motif that consists of approx. 200 amino acids in all eukaryotes. Since the TBC domain of yeast Gyps [GAP (GTPase-activating protein) for Ypt proteins] has been shown to function as a GAP domain for small GTPase Ypt/Rab, TBC domain-containing proteins (TBC proteins) in other species are also expected to function as a certain Rab-GAP. More than 40 different TBC proteins are present in humans and mice, and recent accumulating evidence has indicated that certain mammalian TBC proteins actually function as a specific Rab-GAP. Some mammalian TBC proteins {e.g. TBC1D1 [TBC (Tre-2/Bub2/Cdc16) domain family, member 1] and TBC1D4/AS160 (Akt substrate of 160 kDa)} play an important role in homoeostasis in mammals, and defects in them are directly associated with mouse and human diseases (e.g. leanness in mice and insulin resistance in humans). The present study reviews the structure and function of mammalian TBC proteins, especially in relation to Rab small GTPases.

scholarly journals RASAL3 preferentially stimulates GTP hydrolysis of the Rho family small GTPase Rac2

2018 ◽  
Author(s):  
Yoonjae Shin ◽  
Yong Kim ◽  
Hyemin Kim ◽  
Nakyoung Shin ◽  
Tae Kim ◽  
...  
Keyword(s):  
Small Gtpase ◽  
Gtp Hydrolysis ◽  
Rho Family ◽  
Hydrolysis Of
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