Rap1 up-regulation and activation on plasma membrane regulates T cell adhesion

Rap1 and Ras are closely related GTPases that share some effectors but have distinct functions. We studied the subcellular localization of Rap1 and its sites of activation in living cells. Both GFP-tagged Rap1 and endogenous Rap1 were localized to the plasma membrane (PM) and endosomes. The PM association of GFP-Rap1 was dependent on GTP binding, and GFP-Rap1 was rapidly up-regulated on this compartment in response to mitogens, a process blocked by inhibitors of endosome recycling. A novel fluorescent probe for GTP-bound Rap1 revealed that this GTPase was transiently activated only on the PM of both fibroblasts and T cells. Activation on the PM was blocked by inhibitors of endosome recycling. Moreover, inhibition of endosome recycling blocked the ability of Rap1 to promote integrin-mediated adhesion of T cells. Thus, unlike Ras, the membrane localizations of Rap1 are dynamically regulated, and the PM is the principle platform from which Rap1 signaling emanates. These observations may explain some of the biological differences between these GTPases

PKC Regulates a Farnesyl-Electrostatic Switch on K-Ras that Promotes its Association with Bcl-Xl on Mitochondria and Induces Apoptosis

K-Ras associates with the plasma membrane (PM) through farnesylation that functions in conjunction with an adjacent polybasic sequence. We show that phosphorylation by protein kinase C (PKC) of S181 within the polybasic region promotes rapid dissociation of K-Ras from the PM and association with intracellular membranes, including the outer membrane of mitochondria where phospho-K-Ras interacts with Bcl-Xl. PKC agonists promote apoptosis of cells transformed with oncogenic K-Ras in a S181-dependent manner. K-Ras with a phosphomimetic residue at position 181 induces apoptosis via a pathway that requires Bcl-Xl. The PKC agonist bryostatin-1 inhibited the growth in vitro and in vivo of cells transformed with oncogenic K-Ras in a S181-dependent fashion. These data demonstrate that the location and function of K-Ras are regulated directly by PKC and suggest an approach to therapy of K-Ras-dependent tumors with agents that stimulate phosphorylation of S18

Opening a Pandora's Box that can't be salvaged: Health professionals’ perceptions of appearance-related care in an Australian pediatric specialist hospital

© 2019 Elsevier Ltd Many children and young people struggle adjusting to the psychosocial consequences (e.g., body dissatisfaction, social anxiety, and stigmatisation) of visible differences (or disfigurement). As appearance-affecting conditions often require specialist multidisciplinary team care, health professionals are in a unique position to offer psychosocial support and intervention. However, there is a dearth of literature on how appearance-related concerns are managed in pediatric hospital settings. Sixteen Australian specialist health professionals participated in semi-structured qualitative interviews to address this gap. Interviews explored current appearance-related psychosocial service provision, barriers in accessing appearance-related care, and perceptions of online platforms to deliver specialist support and intervention. Thematic analysis demonstrated four themes: We can do it better, Capability versus availability, Online generation, and Putting appearance on the agenda. This research highlighted the potential value of online platforms to increase accessibility to specialist appearance-related care, the need for more psychosocial resources to be integrated into appearance-related specialities, prioritising the development of low to medium appearance-related support and intervention, increasing the appearance-related knowledge of health professionals and families, and the need for more holistic approaches in routine care

Free movement and EU citizenship: a virtuous circle?

The year 2013 was officially declared the European Year of Citizens (EYC) in the European Union (EU). Through this event, the European Commission (EC) reiterates a ‘virtuous circle’ – between citizenship, free movement and a sense of belonging – able to bring citizens closer to the EU. This article shows how this ‘virtuous circle’ tends to translate into a ‘tunnel vision’ that reduces citizenship to free movement. Through the analysis of EC discourses, of the literature on ‘movers’ and ‘stayers’, and of focus groups with young people from Brussels, we suggest to expand the understanding of free movement and its effects. Overall, this article proposes to re-evaluate the scope of the ‘virtuous circle’ by considering that the ‘stayers’ are also EU citizens, that free movement is not indisputably an attractive right, and that the movers do not unquestionably feel attached to the EU as a result of their mobility

Divergent roles of glycolysis and the mitochondrial electron transport chain in hypoxic pulmonary vasoconstriction of the rat: identity of the hypoxic sensor

The mechanisms responsible for sensing hypoxia and initiating hypoxic pulmonary vasoconstriction (HPV) are unclear. We therefore examined the roles of the mitochondrial electron transport chain (ETC) and glycolysis in HPV of rat small intrapulmonary arteries (IPAs).HPV demonstrated a transient constriction (phase 1) superimposed on a sustained constriction (phase 2). Inhibition of complex I of the ETC with rotenone (100 nm) or complex III with myxothiazol (100 nm) did not cause vasoconstriction in normoxia, but abolished both phases of HPV. Rotenone inhibited the hypoxia-induced rise in intracellular Ca2+ ([Ca2+]i). Succinate (5 mm), a substrate for complex II, reversed the effects of rotenone but not myxothiazol on HPV, but did not affect the rise in NAD(P)H fluorescence induced by hypoxia or rotenone. Inhibition of cytochrome oxidase with cyanide (100 μm) potentiated phase 2 constriction.Phase 2 of HPV, but not phase 1, was highly correlated with glucose concentration, being potentiated by 15 mm but abolished in its absence, or following inhibition of glycolysis by iodoacetate or 2-deoxyglucose. Glucose concentration did not affect the rise in [Ca2+]i during HPV.Depolarisation-induced constriction was unaffected by hypoxia except in the absence of glucose, when it was depressed by ∼50 %. Depolarisation-induced constriction was depressed by rotenone during hypoxia by 23 ± 4 %; cyanide was without effect.Hypoxia increased 2-deoxy-[3H]glucose uptake in endothelium-denuded IPAs by 235 ± 32 %, and in mesenteric arteries by 218 ± 38 %.We conclude that complex III of the mitochondrial ETC acts as the hypoxic sensor in HPV, and initiates the rise in smooth muscle [Ca2+]i by a mechanism unrelated to changes in cytosolic redox state per se, but more probably by increased production of superoxide. Additionally, glucose and glycolysis are essential for development of the sustained phase 2 of HPV, and support an endothelium-dependent Ca2+-sensitisation pathway rather than the rise in [Ca2+]i