C. elegans Rab GTPase activating protein TBC-2 promotes cell corpse degradation by regulating the small GTPase RAB-5

Apoptotic cells generated by programmed cell death are engulfed by phagocytes and enclosed within plasma membrane–derived phagosomes. Maturation of phagosomes involves a series of membrane-remodeling events that are governed by the sequential actions of Rab GTPases and lead to formation of phagolysosomes, where cell corpses are degraded. Here we identified gop-1 as a novel regulator of apoptotic cell clearance in Caenorhabditis elegans. Loss of gop-1 affects phagosome maturation through the RAB-5–positive stage, causing defects in phagosome acidification and phagolysosome formation, phenotypes identical to and unaffected by loss of unc-108, the C. elegans Rab2. GOP-1 transiently associates with cell corpse–containing phagosomes, and loss of its function abrogates phagosomal association of UNC-108. GOP-1 interacts with GDP-bound and nucleotide-free UNC-108/Rab2, disrupts GDI-UNC-108 complexes, and promotes activation and membrane recruitment of UNC-108/Rab2 in vitro. Loss of gop-1 also abolishes association of UNC-108 with endosomes, causing defects in endosome and dense core vesicle maturation. Thus, GOP-1 is an activator of UNC-108/Rab2 in multiple processes.

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The Rab GTPase activating protein TBC-2 extends longevity by facilitating release of FOXO/DAF-16 from endosomes

10.1101/2021.06.04.447106 ◽

2021 ◽

Author(s):

İçten Meraş ◽

Laëtitia Chotard ◽

Thomas Liontis ◽

Zakaria Ratemi ◽

Benjamin Wiles ◽

...

Keyword(s):

Protein Localization ◽

Model Organisms ◽

Rab Gtpase ◽

Gtpase Activating Protein ◽

C Elegans ◽

Functional Consequences ◽

Fat Stores ◽

Foxo Transcription Factors ◽

Endosome Maturation ◽

Insight Into

FOXO transcription factors have been shown to regulate longevity in model organisms and are associated with longevity in humans. To gain insight into how FOXO functions to increase lifespan, we examined the subcellular localization of DAF-16 in C. elegans. We show that DAF-16 is localized to endosomes and that this endosomal localization is increased by the insulin-IGF signaling (IIS) pathway. Endosomal localization of DAF-16 is also increased by disrupting the Rab GTPase activating protein TBC-2, or decreased by inhibiting the RAB-5 or RAB-7 GTPases, key regulators of early to late endosome maturation. Importantly, the amount of DAF-16 that is localized to endosomes has functional consequences as increasing endosomal localization through mutations in tbc-2 decreased the lifespan of long-lived daf-2 IGFR mutants, depleted their fat stores, and DAF-16 target gene expression. Finally, we show that the ability of TBC-2 proteins, TBC1D2 and TBC1D2B, to regulate FOXO protein localization is conserved in human cells. Overall, this work identifies endosomal localization as a mechanism regulating FOXO/DAF-16, which is important for its functions in metabolism and aging.

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TBC proteins: GAPs for mammalian small GTPase Rab?

Bioscience Reports ◽

10.1042/bsr20100112 ◽

2011 ◽

Vol 31 (3) ◽

pp. 159-168 ◽

Cited By ~ 115

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.

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Activation of Cdc42 GTPase upon CRY2-Induced Cortical Recruitment Is Antagonized by GAPs in Fission Yeast

Cells ◽

10.3390/cells9092089 ◽

2020 ◽

Vol 9 (9) ◽

pp. 2089 ◽

Cited By ~ 1

Author(s):

Iker Lamas ◽

Nathalie Weber ◽

Sophie G. Martin

Keyword(s):

Fission Yeast ◽

Positive Feedback ◽

Antagonistic Activity ◽

Cell Polarization ◽

Small Gtpase ◽

Nucleotide Exchange ◽

Gtpase Activating Protein ◽

Guanine Nucleotide Exchange ◽

Nucleotide Exchange Factor ◽

Cell Architecture

The small GTPase Cdc42 is critical for cell polarization in eukaryotic cells. In rod-shaped fission yeast Schizosaccharomyces pombe cells, active GTP-bound Cdc42 promotes polarized growth at cell poles, while inactive Cdc42-GDP localizes ubiquitously also along cell sides. Zones of Cdc42 activity are maintained by positive feedback amplification involving the formation of a complex between Cdc42-GTP, the scaffold Scd2, and the guanine nucleotide exchange factor (GEF) Scd1, which promotes the activation of more Cdc42. Here, we use the CRY2-CIB1 optogenetic system to recruit and cluster a cytosolic Cdc42 variant at the plasma membrane and show that this leads to its moderate activation also on cell sides. Surprisingly, Scd2, which binds Cdc42-GTP, is still recruited to CRY2-Cdc42 clusters at cell sides in individual deletion of the GEFs Scd1 or Gef1. We show that activated Cdc42 clusters at cell sides are able to recruit Scd1, dependent on the scaffold Scd2. However, Cdc42 activity is not amplified by positive feedback and does not lead to morphogenetic changes, due to antagonistic activity of the GTPase activating protein Rga4. Thus, the cell architecture is robust to moderate activation of Cdc42 at cell sides.

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ArhGAP15, a Rac-specific GTPase-activating Protein, Plays a Dual Role in Inhibiting Small GTPase Signaling

Journal of Biological Chemistry ◽

10.1074/jbc.m113.459719 ◽

2013 ◽

Vol 288 (29) ◽

pp. 21117-21125 ◽

Cited By ~ 17

Author(s):

Maria Radu ◽

Sonali J. Rawat ◽

Alexander Beeser ◽

Anton Iliuk ◽

Weiguo Andy Tao ◽

...

Keyword(s):

Catalytic Activity ◽

Protein Microarray ◽

Small Gtpases ◽

Dual Role ◽

Small Gtpase ◽

Ph Domain ◽

Gtpase Activating Protein ◽

Mutual Inhibition ◽

Negative Role ◽

Pak Kinase

Signaling from small GTPases is a tightly regulated process. In this work we used a protein microarray screen to identify the Rac-specific GAP, ArhGAP15, as a substrate of the Rac effectors Pak1 and Pak2. In addition to serving as a substrate of Pak1/2, we found that ArhGAP15, via its PH domain, bound to these kinases. The association of ArhGAP15 to Pak1/2 resulted in mutual inhibition of GAP and kinase catalytic activity, respectively. Knock-down of ArhGAP15 resulted in activation of Pak1/2, both indirectly, as a result of Rac activation, and directly, as a result of disruption of the ArhGAP15/Pak complex. Our data suggest that ArhGAP15 plays a dual negative role in regulating small GTPase signaling, by acting at the level of the GTPase itself, as well interacting with its effector, Pak kinase.

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Measuring Rab GTPase-Activating Protein (GAP) Activity in Live Cells and Extracts

Methods in Molecular Biology - Rab GTPases ◽

10.1007/978-1-4939-2569-8_5 ◽

2015 ◽

pp. 61-71

Author(s):

Ryan M. Nottingham ◽

Suzanne R. Pfeffer

Keyword(s):

Live Cells ◽

Rab Gtpase ◽

Gtpase Activating Protein

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Autophagosomes fuse to phagosomes and play important roles in the degradation of apoptotic cells in Caenorhabditis elegans

10.1101/2021.07.16.452694 ◽

2021 ◽

Author(s):

Omar Pena-Ramos ◽

Lucia Chiao ◽

Xianghua Liu ◽

Tianyou Yao ◽

Henry He ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Small Gtpase ◽

Apoptotic Cells ◽

Phagosome Maturation ◽

Double Membrane ◽

C Elegans ◽

Intracellular Organelles ◽

Intracellular Vesicles ◽

Cellular Components

Autophagosomes are double-membrane intracellular vesicles that degrade protein aggregates, intracellular organelles, and other cellular components. In the nematode Caenorhabditis elegans, 113 somatic cells undergo apoptosis during embryogenesis and are engulfed and degraded by their neighboring cells. We discovered a novel role of autophagosomes in facilitating the degradation of apoptotic cells in C. elegans embryos using a real-time imaging technique. Specifically, double-membrane autophagosomes in engulfing cells are recruited to the surfaces of phagosomes containing apoptotic cells and subsequently fuse to phagosomes, allowing the inner membrane to enter the phagosomal lumen. Mutants defective in the production of autophagosomes display significant delays in the degradation of apoptotic cells, demonstrating the important contribution of autophagosomes to this process. The signaling pathway led by the phagocytic receptor CED-1, CED-1s adaptor CED-6, and the large GTPase dynamin (DYN-1) promote the recruitment of autophagosomes to phagosomes. Moreover, the subsequent fusion of autophagosomes with phagosomes requires the functions of the small GTPase RAB-7 and the HOPS complex. Our findings reveal that, unlike the single-membrane, LC3- associated phagocytosis (LAP) vesicles reported for mammalian phagocytes, canonical autophagosomes function in the clearance of C. elegans apoptotic cells. These findings add autophagosomes to the collection of intracellular organelles that contribute to phagosome maturation, identify novel crosstalk between the autophagy and phagosome maturation pathways, and discover the upstream factors that initiate this crosstalk.

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Three C. elegans Rac proteins and several alternative Rac regulators control axon guidance, cell migration and apoptotic cell phagocytosis

Development ◽

10.1242/dev.128.22.4475 ◽

2001 ◽

Vol 128 (22) ◽

pp. 4475-4488 ◽

Cited By ~ 7

Author(s):

Erik A. Lundquist ◽

Peter W. Reddien ◽

Erika Hartwieg ◽

H. Robert Horvitz ◽

Cornelia I. Bargmann

Keyword(s):

Cell Migration ◽

Axon Guidance ◽

Apoptotic Cell ◽

Regulatory Proteins ◽

Axon Pathfinding ◽

C Elegans ◽

Cell Corpse ◽

And Function ◽

Redundant Functions ◽

Caenorhabditis Elegans Genome

The Caenorhabditis elegans genome contains three rac-like genes, ced-10, mig-2, and rac-2. We report that ced-10, mig-2 and rac-2 act redundantly in axon pathfinding: inactivating one gene had little effect, but inactivating two or more genes perturbed both axon outgrowth and guidance. mig-2 and ced-10 also have redundant functions in some cell migrations. By contrast, ced-10 is uniquely required for cell-corpse phagocytosis, and mig-2 and rac-2 have only subtle roles in this process. Rac activators are also used differentially. The UNC-73 Trio Rac GTP exchange factor affected all Rac pathways in axon pathfinding and cell migration but did not affect cell-corpse phagocytosis. CED-5 DOCK180, which acts with CED-10 Rac in cell-corpse phagocytosis, acted with MIG-2 but not CED-10 in axon pathfinding. Thus, distinct regulatory proteins modulate Rac activation and function in different developmental processes.

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rab-27 acts in an intestinal pathway to inhibit axon regeneration in C. elegans

PLoS Genetics ◽

10.1371/journal.pgen.1009877 ◽

2021 ◽

Vol 17 (11) ◽

pp. e1009877

Author(s):

Alexander T. Lin-Moore ◽

Motunrayo J. Oyeyemi ◽

Marc Hammarlund

Keyword(s):

Motor Neurons ◽

Axon Regeneration ◽

Rab Gtpase ◽

Long Distance ◽

Extrinsic Factors ◽

C Elegans ◽

Cellular Environment ◽

Neuropeptide Secretion ◽

Neuronal Tissues ◽

Nervous System Function

Injured axons must regenerate to restore nervous system function, and regeneration is regulated in part by external factors from non-neuronal tissues. Many of these extrinsic factors act in the immediate cellular environment of the axon to promote or restrict regeneration, but the existence of long-distance signals regulating axon regeneration has not been clear. Here we show that the Rab GTPase rab-27 inhibits regeneration of GABAergic motor neurons in C. elegans through activity in the intestine. Re-expression of RAB-27, but not the closely related RAB-3, in the intestine of rab-27 mutant animals is sufficient to rescue normal regeneration. Several additional components of an intestinal neuropeptide secretion pathway also inhibit axon regeneration, including NPDC1/cab-1, SNAP25/aex-4, KPC3/aex-5, and the neuropeptide NLP-40, and re-expression of these genes in the intestine of mutant animals is sufficient to restore normal regeneration success. Additionally, NPDC1/cab-1 and SNAP25/aex-4 genetically interact with rab-27 in the context of axon regeneration inhibition. Together these data indicate that RAB-27-dependent neuropeptide secretion from the intestine inhibits axon regeneration, and point to distal tissues as potent extrinsic regulators of regeneration.

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