Mussolini, uomo di teatro

Ricerca sullo spettacolo italiano degli anni venti e trenta del novecento

Differential β-arrestin2 requirements for constitutive and agonist-induced internalization of the CB1 cannabinoid receptor

CB1 cannabinoid receptor (CB1R) undergoes both constitutive and agonist-induced internalization, but the underlying mechanisms of these processes and the role of beta-arrestins in the regulation of CB1R function are not completely understood. In this study, we followed CB1R internalization using confocal microscopy and bioluminescence resonance energy transfer measurements in HeLa and Neuro-2a cells. We found that upon activation CB1R binds beta-arrestin2 (beta-arr2), but not beta-arrestin1. Furthermore, both the expression of dominant-negative beta-arr2 (beta-arr2-V54D) and siRNA-mediated knock-down of beta-arr2 impaired the agonist-induced internalization of CB1R. In contrast, neither beta-arr2-V54D nor beta-arr2-specific siRNA had a significant effect on the constitutive internalization of CB1R. However, both constitutive and agonist-induced internalization of CB1R were impaired by siRNA-mediated depletion of clathrin heavy chain. We conclude that although clathrin is required for both constitutive and agonist-stimulated internalization of CB1R, beta-arr2 binding is only required for agonist-induced internalization of the receptor suggesting that the molecular mechanisms underlying constitutive and agonist-induced internalization of CB1R are different

Mutations in the 'DRY' motif of the CB1 cannabinoid receptor result in biased receptor variants.

The role of the highly-conserved 'DRY' motif in the signaling of the CB1 cannabinoid receptor (CB1R) was investigated by introducing single, double and triple alanine mutations into this site of the receptor. We found that the CB1R-R3.50A mutant displays a partial decrease in its ability to activate heterotrimeric Go proteins (~85% of wild-type CB1R (CB1R-WT)). Moreover, this mutant showed impaired beta-arrestin binding in response to agonist stimulus, although its basal beta-arrestin binding was enhanced. More strikingly, the double mutant CB1R-D3.49A/R3.50A was biased toward beta-arrestins, as it gained a robustly increased beta-arrestin1 and beta-arrestin2 binding ability compared to the wild-type receptor, while its G protein activation was decreased. In contrast, the double mutant CB1R-R3.50A/Y3.51A proved to be G protein-biased, as it was practically unable to recruit beta-arrestin2 in response to agonist stimulus, while still activating G proteins, although at a reduced level (~75% of CB1R-WT). Agonist-induced ERK1/2 activation of the CB1R mutants showed good correlation with their beta-arrestin binding ability but not with their G protein activation or inhibition of cAMP accumulation. Our results suggest that G protein-activation and beta-arrestin-binding of the CB1R are mediated by distinct receptor conformations and the conserved 'DRY' motif plays different roles in the stabilization of these conformations, thus mediating both G protein- and beta-arrestin2-mediated functions of CB1R

Effect of a toggle switch mutation in TM6 of the human adenosine A3 receptor on Gi protein-dependent signalling and Gi-independent receptor internalization

Background and Purpose: The highly conserved tryptophan (W6.48) in transmembrane domain 6 of GPCRs has been shown to play a central role in forming an active conformation in response to agonist binding. We set out to characterize the effect of this mutation on the efficacy of two agonists at multiple signalling pathways downstream of the adenosine A3 receptor. Experimental Approach: Residue W6.48 in the human adenosine A3 receptor fused to yellow fluorescent protein was mutated to phenylalanine and expressed in CHO-K1 cells containing a cAMP response element reporter gene. The effects on agonist-mediated receptor internalization were monitored by automated confocal microscopy and image analysis. Further experiments were carried out to investigate agonist-mediated ERK1/2 phosphorylation, inhibition of [3H]-cAMP accumulation and β-arrestin2 binding. Key Results: NECA was able to stimulate agonist-mediated internalization of the W6.48F mutant receptor, while the agonist HEMADO was inactive. Investigation of other downstream signalling pathways indicated that G-protein coupling was impaired for both agonists tested. Mutation of W6.48F therefore resulted in differential effects on agonist efficacy, and introduced signalling pathway bias for HEMADO at the adenosine A3 receptor. Conclusions and Implications: Investigation of the pharmacology of the W6.48F mutant of the adenosine A3 receptor confirms that this region is important in forming the active conformation of the receptor for stimulating a number of different signalling pathways and that mutations in this residue can lead to changes in agonist efficacy and signalling bias

Angiotensin type 1A receptor regulates β-arrestin binding of the β2-adrenergic receptor via heterodimerization

Heterodimerization between angiotensin type 1A receptor (AT1R) and β2-adrenergic receptor (β2AR) has been shown to modulate G protein-mediated effects of these receptors. Activation of G protein-coupled receptors (GPCRs) leads to β-arrestin binding, desensitization, internalization and G protein-independent signaling of GPCRs. Our aim was to study the effect of heterodimerization on β-arrestin coupling. We found that β-arrestin binding of β2AR is affected by activation of AT1Rs. Costimulation with angiotensin II and isoproterenol markedly enhanced the interaction between β2AR and β-arrestins, by prolonging the lifespan of β2AR-induced β-arrestin2 clusters at the plasma membrane. While candesartan, a conventional AT1R antagonist, had no effect on the β-arrestin2 binding to β2AR, TRV120023, a β-arrestin biased agonist, enhanced the interaction. These findings reveal a new crosstalk mechanism between AT1R and β2AR, and suggest that enhanced β-arrestin2 binding to β2AR can contribute to the pharmacological effects of biased AT1R agonists. © 201

Illuminating the life of GPCRs

The investigation of biological systems highly depends on the possibilities that allow scientists to visualize and quantify biomolecules and their related activities in real-time and non-invasively. G-protein coupled receptors represent a family of very dynamic and highly regulated transmembrane proteins that are involved in various important physiological processes. Since their localization is not confined to the cell surface they have been a very attractive "moving target" and the understanding of their intracellular pathways as well as the identified protein-protein-interactions has had implications for therapeutic interventions. Recent and ongoing advances in both the establishment of a variety of labeling methods and the improvement of measuring and analyzing instrumentation, have made fluorescence techniques to an indispensable tool for GPCR imaging. The illumination of their complex life cycle, which includes receptor biosynthesis, membrane targeting, ligand binding, signaling, internalization, recycling and degradation, will provide new insights into the relationship between spatial receptor distribution and function. This review covers the existing technologies to track GPCRs in living cells. Fluorescent ligands, antibodies, auto-fluorescent proteins as well as the evolving technologies for chemical labeling with peptide- and protein-tags are described and their major applications concerning the GPCR life cycle are presented

Podatność na gięcie drewna warstwowego modyfikowanego tworzywami sztucznymi

The bending properties of laminated wood combined with plastic. In the article we present the bending properties of laminated wood produced by the combination of beech veneer and plastic. The combination of wood and plastic enables us to create materials with different properties compared to solid wood. A small decrease in bending strength was demonstrated greater bendability laminated timber.Podatność na gięcie drewna warstwowego modyfikowanego tworzywami sztucznymi. Praca opisuje podatność na gięcie drewna warstwowego powstałego przez spajanie fornirów bukowych oraz plastiku. Taki kompozyt ma własności mechaniczne różne od własności jego poszczególnych składników, testy wykazały większą podatność na gięcie przy nieco zmniejszonej wytrzymałości kompozytu, w porównaniu do drewna litego

Brancati’s Gigantic Mussolinis

Best known for such novels as Il bell’Antonio and Paolo il Caldo, Sicilian writer Vitaliano Brancati (Pachino, 1907) was also a playwright and, in his younger years, so enthused about dictator Benito Mussolini that he was a propagandist for the fascist regime. This article explores his figuration of Mussolini as he appears in three of the dramatic works written in this period: Everest (1928), Piave (1932), and L’Urto (1934). The discussion focuses on Brancati’s depiction of the “Duce” as “larger than life” – a metaphor that becomes literal in the plays’ characterizations of the dictator – to address, first, the ways in which Brancati’s vision of Mussolini intersects with the regime’s mythologization and sacralization of the leader, and, second, how the plays shed light on fascist intellectuals’ ambitions to create propaganda art for the would-be new era

Wpływ grubości na możliwości gięcia drewna osiki

Influence of thickness on the bendability of aspen wood. Aspen is a little industrial exploitation of wood. For its application in the manufacture of molded furniture, it is necessary to know its bendability. In this work we focused on the influence of the thickness on the bendability of aspen wood. Bendability varies depending on the thickness.Wpływ grubości na możliwości gicia drewna osiki. Osika ma niewielkie zastosowanie w przemyśle. Aby opracować założenia technologiczne jego zastosowania w meblach giętych, należy zbadać możliwość gięcia tego drewna. W pracy skupiono się na możliwości gięcia drewna osiki w zależności od jego grubości, wykazano że te możliwości zmieniają się wraz z grubością