Lizofoszfolipid mediátorok receptorai. Szfingo- és glicerolipid növekedési faktorokat kötő fehérjék azonosítása és jellemzése. = Identification and characterization of specific proteins for sphingo- and glycerolipid growth factors

A legegyszerűbb foszfolipidek, a lizofoszfatidsav (LPA) és a szfingozin-1-foszfát (S1P) jelátvivő molekulák, amelyek részt vesznek a sejtek túlélésének, osztódásának és mozgásainak szabályozásában az egyszerű organizmusoktól az emberig. Kutatásaink célja ezen foszfolipidek hatásmechanizmusának megismerése, a kölcsönható fehérjék azonosítása. Az LPA és az S1P hatását elsősorban sejtfelszíni receptorokon keresztül fejti ki. Az LPA-receptortípusokra szelektív aktiváló és gátló molekulák két új csoportját azonosítottuk és jellemeztük: a zsíralkohol-foszfátokat és az oligoprenil-foszfátokat. Sikeresen azonosítottuk az S1P1-es típusú receptor ligandkötésében résztvevő apoláros aminosavakat, teljessé téve ezzel korábbi munkánkat a kötőhely feltérképezésére. Eredményeink a lizofoszfolipid receptorok farmakológiai vizsgálatának új lehetőségeit teremtik meg. Az S1P különlegessége, hogy másodlagos hírvivő is. Egyértelmű bizonyítékot szolgáltattunk arra, hogy az S1P valóban intracellulárisan is képes a kalcium-ionok felszabadítására. Legújabban kimutattuk, hogy az S1P-rokon szfingozilfoszforilkolin köti és gátolja a kalcium hatását univerzálisan közvetítő kalmodulint. Az S1P a sejtben keletkezik, tehát sejtfelszíni receptorai aktiválásához ki kell jutnia onnan. Vizsgáltuk egyes ABC-transzporterek szerepét ebben a folyamatban. Kimutattuk, hogy az MRP1-fehérje az S1P egyik lehetséges kipumpálója. E fehérje működése során fellépő kooperatív kölcsönhatásokat is azonosítottunk. | The simplest phospholipids, lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) are lipid mediators regulating survival, proliferation, and migration of cells. Our aim is to characterize the mechanisms of action of these phospholipids and to identify their interacting proteins. LPA and S1P exert their actions mainly through cell surface G protein-coupled receptors. We identified two new sets of LPA receptor subtype selective ligands: fatty alcohol phosphates and oligoprenyl phosphates. We also successfully completed the mapping of the ligand binding pocket of the S1P1 receptor, by determining the amino acids lining the hydrophobic part of the binding site. Our results make new pharmacologic interventions possible at lysophospholipid receptors. S1P is unique because it also acts as a second messenger. We provided strict evidence for the action of S1P on the intracellular calcium mobilization from endoplasmic reticulum, independent of cell surface receptors. Very recently we have shown that the related lipid sphingosylphosphorylcholine binds to and inhibits the actions of calmodulin, the ubiquitous calcium sensor of cells. S1P is formed inside cells by the action of sphingosine kinases. It should leave the cell to act as an autocrine/paracrine mediator. We investigated the role of ABC transporters in this process and identified MRP1 as a potential efflux pump for S1P. We also identified cooperative interactions between the ATP and drug binding sites of MRP1

Downregulation of sphingosine 1-phosphate (S1P) receptor 1 by dexamethasone inhibits S1P-induced mesangial cell migration

Sphingosine 1-phosphate (S1P) is generated by sphingosine kinase (SK)-1 and -2 and acts mainly as an extracellular ligand at five specific receptors, denoted S1P1-5. After activation, S1P receptors regulate important processes in the progression of renal diseases, such as mesangial cell migration and survival. Previously, we showed that dexamethasone enhances SK-1 activity and S1P formation, which protected mesangial cells from stress-induced apoptosis. Here we demonstrate that dexamethasone treatment lowered S1P1 mRNA and protein expression levels in rat mesangial cells. This effect was abolished in the presence of the glucocorticoid receptor antagonist RU-486. In addition, in vivo studies showed that dexamethasone downregulated S1P1 expression in glomeruli isolated from mice treated with dexamethasone (10 mg/kg body weight). Functionally, we identified S1P1 as a key player mediating S1P-induced mesangial cell migration. We show that dexamethasone treatment significantly lowered S1P-induced migration of mesangial cells, which was again reversed in the presence of RU-486. In summary, we suggest that dexamethasone inhibits S1P-induced mesangial cell migration via downregulation of S1P1. Overall, these results demonstrate that dexamethasone has functional important effects on sphingolipid metabolism and action in renal mesangial cells

Downregulation of sphingosine 1-phosphate (S1P) receptor 1 by dexamethasone inhibits S1P-induced mesangial cell migration

Sphingosine 1-phosphate (S1P) is generated by sphingosine kinase (SK)-1 and -2 and acts mainly as an extracellular ligand at five specific receptors, denoted S1P1-5. After activation, S1P receptors regulate important processes in the progression of renal diseases, such as mesangial cell migration and survival. Previously, we showed that dexamethasone enhances SK-1 activity and S1P formation, which protected mesangial cells from stress-induced apoptosis. Here we demonstrate that dexamethasone treatment lowered S1P1 mRNA and protein expression levels in rat mesangial cells. This effect was abolished in the presence of the glucocorticoid receptor antagonist RU-486. In addition, in vivo studies showed that dexamethasone downregulated S1P1 expression in glomeruli isolated from mice treated with dexamethasone (10 mg/kg body weight). Functionally, we identified S1P1 as a key player mediating S1P-induced mesangial cell migration. We show that dexamethasone treatment significantly lowered S1P-induced migration of mesangial cells, which was again reversed in the presence of RU-486. In summary, we suggest that dexamethasone inhibits S1P-induced mesangial cell migration via downregulation of S1P1. Overall, these results demonstrate that dexamethasone has functional important effects on sphingolipid metabolism and action in renal mesangial cells

Mouse Liver Compensates Loss of Sgpl1 by Secretion of Sphingolipids into Blood and Bile

Sphingosine 1 phosphate (S1P) lyase (Sgpl1) catalyses the irreversible cleavage of S1P and thereby the last step of sphingolipid degradation. Loss of Sgpl1 in humans and mice leads to accumulation of sphingolipids and multiple organ injuries. Here, we addressed the role of hepatocyte Sgpl1 for regulation of sphingolipid homoeostasis by generating mice with hepatocyte-specific deletion of Sgpl1 (Sgpl1HepKO mice). Sgpl1HepKO mice had normal body weight, liver weight, liver structure and liver enzymes both at the age of 8 weeks and 8 months. S1P, sphingosine and ceramides, but not glucosylceramides or sphingomyelin, were elevated by ~1.5–2-fold in liver, and this phenotype did not progress with age. Several ceramides were elevated in plasma, while plasma S1P was normal. Interestingly, S1P and glucosylceramides, but not ceramides, were elevated in bile of Sgpl1HepKO mice. Furthermore, liver cholesterol was elevated, while LDL cholesterol decreased in 8-month-old mice. In agreement, the LDL receptor was upregulated, suggesting enhanced uptake of LDL cholesterol. Expression of peroxisome proliferator-activated receptor-γ, liver X receptor and fatty acid synthase was unaltered. These data show that mouse hepatocytes largely compensate the loss of Sgpl1 by secretion of accumulating sphingolipids in a specific manner into blood and bile, so that they can be excreted or degraded elsewher

Downregulation of sphingosine 1-phosphate (S1P) receptor 1 by dexamethasone inhibits S1P-induced mesangial cell migration

Sphingosine 1-phosphate (S1P) is generated by sphingosine kinase (SK)-1 and -2 and acts mainly as an extracellular ligand at five specific receptors, denoted S1P1-5. After activation, S1P receptors regulate important processes in the progression of renal diseases, such as mesangial cell migration and survival. Previously, we showed that dexamethasone enhances SK-1 activity and S1P formation, which protected mesangial cells from stress-induced apoptosis. Here we demonstrate that dexamethasone treatment lowered S1P1 mRNA and protein expression levels in rat mesangial cells. This effect was abolished in the presence of the glucocorticoid receptor antagonist RU-486. In addition, in vivo studies showed that dexamethasone downregulated S1P1 expression in glomeruli isolated from mice treated with dexamethasone (10 mg/kg body weight). Functionally, we identified S1P1 as a key player mediating S1P-induced mesangial cell migration. We show that dexamethasone treatment significantly lowered S1P-induced migration of mesangial cells, which was again reversed in the presence of RU-486. In summary, we suggest that dexamethasone inhibits S1P-induced mesangial cell migration via downregulation of S1P1. Overall, these results demonstrate that dexamethasone has functional important effects on sphingolipid metabolism and action in renal mesangial cells

Sphingosine 1-phosphate (S1P) induces COX-2 expression and PGE2 formation via S1P receptor 2 in renal mesangial cells

Understanding the mechanisms of sphingosine 1-phosphate (S1P)-induced cyclooxygenase (COX)-2 expression and prostaglandin E2 (PGE2) formation in renal mesangial cells may provide potential therapeutic targets to treat inflammatory glomerular diseases. Thus, we evaluated the S1P-dependent signaling mechanisms which are responsible for enhanced COX-2 expression and PGE2 formation in rat mesangial cells under basal conditions. Furthermore, we investigated whether these mechanisms are operative in the presence of angiotensin II (Ang II) and of the pro-inflammatory cytokine interleukin-1β (IL-1β). Treatment of rat and human mesangial cells with S1P led to concentration-dependent enhanced expression of COX-2. Pharmacological and molecular biology approaches revealed that the S1P-dependent increase of COX-2 mRNA and protein expression was mediated via activation of S1P receptor 2 (S1P2). Further, inhibition of Gi and p42/p44 MAPK signaling, both downstream of S1P2, abolished the S1P-induced COX-2 expression. In addition, S1P/S1P2-dependent upregulation of COX-2 led to significantly elevated PGE2 levels, which were further potentiated in the presence of Ang II and IL-1β. A functional consequence downstream of S1P/S1P2 signaling is mesangial cell migration that is stimulated by S1P. Interestingly, inhibition of COX-2 by celecoxib and SC-236 completely abolished the migratory response. Overall, our results demonstrate that extracellular S1P induces COX-2 expression via activation of S1P2 and subsequent Gi and p42/p44 MAPK-dependent signaling in renal mesangial cells leading to enhanced PGE2 formation and cell migration that essentially requires COX-2. Thus, targeting S1P/S1P2 signaling pathways might be a novel strategy to treat renal inflammatory diseases

cAMP guided his way: a life for G protein-mediated signal transduction and molecular pharmacology-tribute to Karl H. Jakobs

Karl H. Jakobs, former editor-in-chief of Naunyn-Schmiedeberg's Archives of Pharmacology and renowned molecular pharmacologist, passed away in April 2018. In this article, his scientific achievements regarding G protein-mediated signal transduction and regulation of canonical pathways are summarized. Particularly, the discovery of inhibitory G proteins for adenylyl cyclase, methods for the analysis of receptor-G protein interactions, GTP supply by nucleoside diphosphate kinases, mechanisms in phospholipase C and phospholipase D activity regulation, as well as the development of the concept of sphingosine-1-phosphate as extra- and intracellular messenger will presented. His seminal scientific and methodological contributions are put in a general and timely perspective to display and honor his outstanding input to the current knowledge in molecular pharmacology