Santa Maria Imbaro, Italy
Santa Maria Imbaro, Italy

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Dani N.,G Protein mediated Signalling Laboratory | Mayo E.,G Protein mediated Signalling Laboratory | Stilla A.,G Protein mediated Signalling Laboratory | Marchegiani A.,G Protein mediated Signalling Laboratory | And 4 more authors.
Journal of Biological Chemistry | Year: 2011

Mono-ADP-ribosylation is a reversible post-translational modification that can modulate the functions of target proteins. We have previously demonstrated that the β subunit of heterotrimeric G proteins is endogenously mono-ADP-ribosylated, and once modified, the βγ dimer is inactive toward its effector enzymes. To better understand the physiological relevance of this post-translational modification, we have studied its hormonal regulation. Here, we report that Gβ subunit mono-ADP-ribosylation is differentially modulated by G protein-coupled receptors. In intact cells, hormone stimulation of the thrombin receptor induces Gβ subunit mono-ADP-ribosylation, which can affect G protein signaling. Conversely, hormone stimulation of the gonadotropin-releasing hormone receptor (GnRHR) inhibits Gβ subunit mono-ADP-ribosylation. We also provide the first demonstration that activation of the GnRHR can activate the ADP-ribosylation factor Arf6, which in turn inhibits Gβ subunit mono-ADP-ribosylation. Indeed, removal of Arf6 from purified plasma membranes results in loss of GnRHR-mediated inhibition of Gβ subunit mono-ADP-ribosylation, which is fully restored by re-addition of purified, myristoylated Arf6. We show that Arf6 acts as a competitive inhibitor of the endogenous ADP-ribosyltransferase and is itself modified by this enzyme. These data provide further understanding of the mechanisms that regulate endogenous ADP-ribosylation of the Gβ subunit, and they demonstrate a novel role for Arf6 in hormone regulation of Gβ subunit mono-ADP-ribosylation. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.


Di Girolamo M.,G Protein Mediated Signalling Laboratory
Current Topics in Microbiology and Immunology | Year: 2014

Post-translational modifications of a cellular protein by mono- and poly- ADP-ribosylation involve the cleavage of NAD+, with the release of its nicotinamide moiety. This is accompanied by the transfer of a single (mono-) or several (poly-) ADP-ribose molecules from NAD+ to a specific amino-acid residue of the protein. Recent reports have shed new light on the correlation between NAD+- dependent ADP-ribosylation reactions and the endoplasmic reticulum, in addition to the well-documented roles of these reactions in the nucleus and mitochondria. We have demonstrated that ARTD15/PARP16 is a novel mono-ADP-ribosyltransferase with a new intracellular location, as it is associated with the endoplasmic reticulum. The endoplasmic reticulum, which is a membranous network of interconnected tubules and cisternae, is responsible for specialised cellular functions, including protein folding and protein transport. Maintenance of specialised cellular functions requires the correct flow of information between separate organelles that is made possible through the nucleocytoplasmic trafficking of proteins. ARTD15 appears to have a role in nucleocytoplasmic shuttling, through karyopherin-β1 mono- ADP-ribosylation. This is in line with the emerging role of ADP-ribosylation in the regulation of intracellular trafficking of cellular proteins. Indeed, other,ADPribosyltransferases like ARTD1/PARP1, have been reported to regulate nucleocytoplasmic trafficking of crucial proteins, including p53 and NF-KB, and as a consequence, to modulate the subcellular localisation of these proteins under both physiological and pathological conditions. © Springer International Publishing Switzerland 2014.


PubMed | G Protein mediated Signalling Laboratory
Type: Journal Article | Journal: The Journal of biological chemistry | Year: 2011

Mono-ADP-ribosylation is a reversible post-translational modification that can modulate the functions of target proteins. We have previously demonstrated that the subunit of heterotrimeric G proteins is endogenously mono-ADP-ribosylated, and once modified, the dimer is inactive toward its effector enzymes. To better understand the physiological relevance of this post-translational modification, we have studied its hormonal regulation. Here, we report that G subunit mono-ADP-ribosylation is differentially modulated by G protein-coupled receptors. In intact cells, hormone stimulation of the thrombin receptor induces G subunit mono-ADP-ribosylation, which can affect G protein signaling. Conversely, hormone stimulation of the gonadotropin-releasing hormone receptor (GnRHR) inhibits G subunit mono-ADP-ribosylation. We also provide the first demonstration that activation of the GnRHR can activate the ADP-ribosylation factor Arf6, which in turn inhibits G subunit mono-ADP-ribosylation. Indeed, removal of Arf6 from purified plasma membranes results in loss of GnRHR-mediated inhibition of G subunit mono-ADP-ribosylation, which is fully restored by re-addition of purified, myristoylated Arf6. We show that Arf6 acts as a competitive inhibitor of the endogenous ADP-ribosyltransferase and is itself modified by this enzyme. These data provide further understanding of the mechanisms that regulate endogenous ADP-ribosylation of the G subunit, and they demonstrate a novel role for Arf6 in hormone regulation of G subunit mono-ADP-ribosylation.

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