PAF Receptor Trafficking and Function in Epithelial Cells

Summary

Principal Investigator: Tamas Jilling
Abstract: Platelet-activating factor (PAF) has been implicated as a key mediator in the pathogenesis of gastrointestinal diseases such as Crohn's disease, ulcerative colitis and neonatal necrotizing enterocolitis (NEC). The PAF receptor (PAFR) is a member of the G protein-coupled receptor (GPCR) superfamily. Despite the importance of PAF in gastrointestinal diseases, and the high level expression of PAFR in intestinal epithelial cells, PAFR trafficking and signal transduction have been studied only in non-epithelial tissues. Our previous studies have shown that PAF plays a crucial role in experimental NEC, PAFR is localized exclusively in the apical plasma membrane in cultured intestinal epithelial cells and regulates such diverse cellular functions as gene expression, ion transport, intracellular pH, and cell death. Preliminary data presented in this proposal shows that disruption of detergent resistant membrane domains (DRM), blocking palmitoylation, or treatment of intestinal epithelial cells with polyunsaturated fatty acids (PUFA) eliminates, or blunts PAF-induced cellular responses. PUFA are important nutrients, with a broad range of biological effects in development, health and disease. In particular, PUFA play a preventive role in an experimental model of NEC, where PAF is a critical mediator. We hypothesize that PAFR is targeted to DRM via palmitoylation of C317 in its cytoplasmic tail, and that disruption of palmitoylation and DRM targeting by PUFA can inhibit the efficiency of signal transduction by PAFR. This hypothesis will be tested based on the following three specific aims: 1) To identify the mechanisms that target PAFR to DRM in the apical plasma membrane of polarized epithelial cells and to determine the importance of DRM targeting in PAFR signaling. 2) To determine whether PUFA can modulate signaling by affecting PAFR targeting to DRM. 3) To examine the effect of PAFR palmitoylation and PUFA on PAFR targeting and on experimental NEC in vivo. In order to accomplish these goals we will utilize heterologously expressed, tagged wild type and mutant PAFR in polarized epithelial cell lines, in neonatal rat intestine using adenoviral gene transfer and in transgenic mice along with the pharmacological manipulation of DRM and palmitoylation. Targeting of PAFR will be analyzed using imaging, immunologic and biochemical methods, and function will be evaluated using highly reproducible functional assays. These studies will elucidate the mechanisms of PAFR targeting in epithelial cells, and a novel mechanism by which PUFA might affect the function of PAFR, and other GPCR-s.
Funding Period: ----------------2006 - ---------------2011-
more information: NIH RePORT

Top Publications

  1. pmc The roles of bacteria and TLR4 in rat and murine models of necrotizing enterocolitis
    Tamas Jilling
    Department of Pediatrics, Northwestern University Feinberg School of Medicine, Chicago, IL 60614, USA
    J Immunol 177:3273-82. 2006
  2. ncbi Inhibition of nuclear factor-kappaB ameliorates bowel injury and prolongs survival in a neonatal rat model of necrotizing enterocolitis
    Isabelle G De Plaen
    Department of Pediatrics, Children s Memorial Hospital, Northwestern University Medical School, Chicago, Illinois 60614, USA
    Pediatr Res 61:716-21. 2007
  3. pmc Polyunsaturated fatty acid supplementation alters proinflammatory gene expression and reduces the incidence of necrotizing enterocolitis in a neonatal rat model
    Jing Lu
    Evanston Northwestern Healthcare Research Institute, Evanston, IL 60201, USA
    Pediatr Res 61:427-32. 2007
  4. pmc Platelet-activating factor-induced chloride channel activation is associated with intracellular acidosis and apoptosis of intestinal epithelial cells
    Erika C Claud
    Department of Pediatrics, University of Chicago, Chicago, IL 60637, USA
    Am J Physiol Gastrointest Liver Physiol 294:G1191-200. 2008
  5. pmc The importance of pro-inflammatory signaling in neonatal necrotizing enterocolitis
    Brandy L Frost
    Department of Pediatrics, Evanston Northwestern Healthcare Feinberg School of Medicine, Northwestern University, Evanston, IL 60201, USA
    Semin Perinatol 32:100-6. 2008
  6. pmc Polyunsaturated fatty acids block platelet-activating factor-induced phosphatidylinositol 3 kinase/Akt-mediated apoptosis in intestinal epithelial cells
    Jing Lu
    Evanston Northwestern Healthcare Research Institute, Evanston, IL 60201, USA
    Am J Physiol Gastrointest Liver Physiol 294:G1181-90. 2008
  7. pmc Dual roles of endogenous platelet-activating factor acetylhydrolase in a murine model of necrotizing enterocolitis
    Jing Lu
    Department of Pediatrics, Pritzker School of Medicine, University of Chicago, Evanston, IL 60201, USA
    Pediatr Res 68:225-30. 2010
  8. pmc Platelet-activating factor induces TLR4 expression in intestinal epithelial cells: implication for the pathogenesis of necrotizing enterocolitis
    Antoine Soliman
    Department of Pediatrics, University of California Los Angeles School of Medicine, Los Angeles, California, United States of America
    PLoS ONE 5:e15044. 2010

Scientific Experts

  • Moshe Arditi
  • Jing Lu
  • Tamas Jilling
  • Michael S Caplan
  • Antoine Soliman
  • Dan Li
  • Erika C Claud
  • Brandy L Frost
  • Fan Jing Meng
  • Isabelle G De Plaen
  • Xiaowu Qu
  • Kathrin S Michelsen
  • Marissa Pierce
  • Diana M Stafforini
  • Andrew Franklin
  • Shuang Chen
  • Michael Caplan
  • SHERVIN RABIZADEH
  • Hisae Karahashi
  • Timothy R Crother
  • Deborah J Nelson
  • Elaine O Petrof
  • Jeremy Marks
  • Xue Qing Wang
  • Jun Sun
  • Mark Abe
  • Wei Hsueh
  • Xin Bing Han
  • Isaac Neequaye
  • Runlan Tian
  • Michael J May
  • Shirley X L Liu
  • Dyan Simon
  • Robert Schy
  • Richard B Thomson

Detail Information

Publications9

  1. pmc The roles of bacteria and TLR4 in rat and murine models of necrotizing enterocolitis
    Tamas Jilling
    Department of Pediatrics, Northwestern University Feinberg School of Medicine, Chicago, IL 60614, USA
    J Immunol 177:3273-82. 2006
    ..In summary, bacteria and TLR4 play significant roles in experimental NEC, likely via an interaction of intraluminal bacteria and aberrantly overexpressed TLR4 in enterocytes...
  2. ncbi Inhibition of nuclear factor-kappaB ameliorates bowel injury and prolongs survival in a neonatal rat model of necrotizing enterocolitis
    Isabelle G De Plaen
    Department of Pediatrics, Children s Memorial Hospital, Northwestern University Medical School, Chicago, Illinois 60614, USA
    Pediatr Res 61:716-21. 2007
    ..Our findings therefore lead us to conclude that selective NF-kappaB inhibition represents a promising therapeutic strategy for NEC...
  3. pmc Polyunsaturated fatty acid supplementation alters proinflammatory gene expression and reduces the incidence of necrotizing enterocolitis in a neonatal rat model
    Jing Lu
    Evanston Northwestern Healthcare Research Institute, Evanston, IL 60201, USA
    Pediatr Res 61:427-32. 2007
    ..These results suggest that PUFA modulates gene expression of key factors involved in experimental NEC pathogenesis. These effects might in part explain the protective effect of PUFA on neonatal NEC...
  4. pmc Platelet-activating factor-induced chloride channel activation is associated with intracellular acidosis and apoptosis of intestinal epithelial cells
    Erika C Claud
    Department of Pediatrics, University of Chicago, Chicago, IL 60637, USA
    Am J Physiol Gastrointest Liver Physiol 294:G1191-200. 2008
    ..Our data suggest a novel mechanism of PAF-induced injury by which PAF induces intracellular acidosis via activation of the Ca(2+)-dependent Cl(-) channel ClC-3, resulting in apoptosis of IEC...
  5. pmc The importance of pro-inflammatory signaling in neonatal necrotizing enterocolitis
    Brandy L Frost
    Department of Pediatrics, Evanston Northwestern Healthcare Feinberg School of Medicine, Northwestern University, Evanston, IL 60201, USA
    Semin Perinatol 32:100-6. 2008
    ..In this review, we examine the key points in the signaling pathways involved in NEC, and potential strategies for prevention and treatment of this dreaded disease...
  6. pmc Polyunsaturated fatty acids block platelet-activating factor-induced phosphatidylinositol 3 kinase/Akt-mediated apoptosis in intestinal epithelial cells
    Jing Lu
    Evanston Northwestern Healthcare Research Institute, Evanston, IL 60201, USA
    Am J Physiol Gastrointest Liver Physiol 294:G1181-90. 2008
    ....
  7. pmc Dual roles of endogenous platelet-activating factor acetylhydrolase in a murine model of necrotizing enterocolitis
    Jing Lu
    Department of Pediatrics, Pritzker School of Medicine, University of Chicago, Evanston, IL 60201, USA
    Pediatr Res 68:225-30. 2010
    ..In conclusion, the data support a protective role for endogenous PAF-AH in the development of NEC, and because preterm neonates have endogenous PAF-AH deficiency, this may place them at increased risk for disease...
  8. pmc Platelet-activating factor induces TLR4 expression in intestinal epithelial cells: implication for the pathogenesis of necrotizing enterocolitis
    Antoine Soliman
    Department of Pediatrics, University of California Los Angeles School of Medicine, Los Angeles, California, United States of America
    PLoS ONE 5:e15044. 2010
    ..Our findings provide evidence for a mechanism by which PAF augments inflammation in the intestinal epithelium through abnormal TLR4 upregulation, thereby contributing to the intestinal injury of NEC...