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ADP-ribosylation of the GTP-binding protein Rho by Clostridium limosum exoenzyme affects basal, but not N-formyl-peptide-stimulated, actin polymerization in human myeloid leukaemic (HL60) cells.

ABSTRACT: Treatment of human myeloid leukaemic (HL60) cells with Clostridium limosum exoenzyme, which inactivates the small GTP-binding protein Rho by ADP-ribosylation, decreased the basal F-actin content. Inhibition of F-actin occurred after long-term treatment (24 h) of intact HL60 cells or after introduction of the toxin by electropermeabilization in a toxin-concentration-dependent manner. Concomitantly with the decrease in the basal F-actin content, the GTP-binding protein Rho was ADP-ribosylated in intact cells. However, Clostridium limosum toxin had no inhibitory effect on N-formyl-peptide-induced actin polymerization. Moreover, the relative N-formyl-peptide-stimulated polymerization was substantially enhanced in cells treated with Clostridium limosum transferase. In contrast with Clostridium limosum exoenzyme, component C21 of the Clostridium botulinum C2 toxin, which ADP-ribosylates G-actin, depolymerized basal F-actin and inhibited N-formyl-peptide-induced actin polymerization in electropermeabilized HL60 cells. These findings indicate that Rho proteins are involved in the basal, but not the ligand-evoked, actin polymerization in HL60 cells.

SUBMITTER: Koch G

PROVIDER: S-EPMC1138088 | biostudies-other | 1994 May

REPOSITORIES: biostudies-other

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ADP-ribosylation of Rho proteins by Clostridium botulinum exoenzyme C3 is influenced by phosphorylation of Rho-associated factors.

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The Rho ADP-ribosylating C3 exoenzyme binds cells via an Arg-Gly-Asp motif.

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Structural basis of actin recognition and arginine ADP-ribosylation by Clostridium perfringens iota-toxin.

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Roles of Asp179 and Glu270 in ADP-Ribosylation of Actin by Clostridium perfringens Iota Toxin.

Project description:Clostridium perfringens iota toxin is a binary toxin composed of the enzymatically active component Ia and receptor binding component Ib. Ia is an ADP-ribosyltransferase, which modifies Arg177 of actin. The previously determined crystal structure of the actin-Ia complex suggested involvement of Asp179 of actin in the ADP-ribosylation reaction. To gain more insights into the structural requirements of actin to serve as a substrate for toxin-catalyzed ADP-ribosylation, we engineered Saccharomyces cerevisiae strains, in which wild type actin was replaced by actin variants with substitutions in residues located on the Ia-actin interface. Expression of the actin mutant Arg177Lys resulted in complete resistance towards Ia. Actin mutation of Asp179 did not change Ia-induced ADP-ribosylation and growth inhibition of S. cerevisiae. By contrast, substitution of Glu270 of actin inhibited the toxic action of Ia and the ADP-ribosylation of actin. In vitro transcribed/translated human ?-actin confirmed the crucial role of Glu270 in ADP-ribosylation of actin by Ia.

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Reversible ADP-ribosylation of RNA.

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Proteasome regulation by ADP-ribosylation.

Project description:Protein degradation by the ubiquitin-proteasome system is central to cell homeostasis and survival. Defects in this process are associated with diseases such as cancer and neurodegenerative disorders. The 26S proteasome is a large protease complex that degrades ubiquitinated proteins. Here, we show that ADP-ribosylation promotes 26S proteasome activity in both Drosophila and human cells. We identify the ADP-ribosyltransferase tankyrase (TNKS) and the 19S assembly chaperones dp27 and dS5b as direct binding partners of the proteasome regulator PI31. TNKS-mediated ADP-ribosylation of PI31 drastically reduces its affinity for 20S proteasome α subunits to relieve 20S repression by PI31. Additionally, PI31 modification increases binding to and sequestration of dp27 and dS5b from 19S regulatory particles, promoting 26S assembly. Inhibition of TNKS by either RNAi or a small-molecule inhibitor, XAV939, blocks this process to reduce 26S assembly. These results unravel a mechanism of proteasome regulation that can be targeted with existing small-molecule inhibitors.

| S-EPMC3676968 | biostudies-literature

Exoenzyme S shows selective ADP-ribosylation and GTPase-activating protein (GAP) activities towards small GTPases in vivo.

Project description:Intracellular targeting of the Pseudomonas aeruginosa toxins exoenzyme S (ExoS) and exoenzyme T (ExoT) initially results in disruption of the actin microfilament structure of eukaryotic cells. ExoS and ExoT are bifunctional cytotoxins, with N-terminal GTPase-activating protein (GAP) and C-terminal ADP-ribosyltransferase activities. We show that ExoS can modify multiple GTPases of the Ras superfamily in vivo. In contrast, ExoT shows no ADP-ribosylation activity towards any of the GTPases tested in vivo. We further examined ExoS targets in vivo and observed that ExoS modulates the activity of several of these small GTP-binding proteins, such as Ras, Rap1, Rap2, Ral, Rac1, RhoA and Cdc42. We suggest that ExoS is the major ADP-ribosyltransferase protein modulating small GTPase function encoded by P. aeruginosa. Furthermore, we show that the GAP activity of ExoS abrogates the activation of RhoA, Cdc42 and Rap1.

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