TLR-mediated Signaling Complex Formation and Regulation of Effector Functions

Summary

Principal Investigator: Hans Hacker
Affiliation: St. Jude Children's Research Hospital
Country: USA
Abstract: The major goal of this project is to define and characterize unknown components of the signaling pathways triggered by Toll-like receptors (TLRs). The function of TLRs on innate immune cells is to recognize pathogens and translate recognition into cell activation and appropriate effector functions, e.g. cytokine production. Ten TLRs are known in humans that specifically recognize diverse pathogen constituents such as lipopolysaccharides (LPS), peptidoglycans (PGN), nucleotides and proteins. This enormous range of specificities enables innate immune cells to respond immediately to virtually all classes of pathogens, including bacteria, fungi, viruses and protozoans. In addition to their physiological role in immune defense, TLRs are also critically involved in the development of immune pathology as elicited by inappropriate TLR stimulation during sepsis and autoimmune diseases. As such, the TLRs and their signal transduction pathways represent important targets for therapeutic intervention strategies. Several hierarchically acting key components of the TLR signal transduction pathways have been defined: TLR activation leads to recruitment and oligomerization of the adaptor protein MyD88, which binds to members of the IRAK family, which in turn recruit TRAF6. TRAF6 oligomerization induces diverse signaling pathways, eventually leading to activation of downstream effector kinases that directly activate transcription factors. Still, it is unknown whether additional proteins exist that act in between MyD88 and TRAF6. Also, little is known about the signaling events between TRAF6 and many of the downstream effector kinases. One reason for this lack of information is that it is technically difficult to explore transiently assembled signaling complexes. We have developed a robust and sensitive technique that allows purification and characterization of such signaling complexes. In this procedure, TLR-mediated dimerization of signaling proteins is mimicked by fusion of these proteins to the bacterial protein Gyrase B, which can inducibly be dimerized by the bivalent antibiotic coumermycin A1. When fused to MyD88 or TRAF6 and equipped with additional epitope tags, the highly selective signaling complexes can be purified and characterized by mass spectrometry (MS). We have demonstrated the proof of principle for this approach by identifying TRAF3 as a critical component of TLR signaling pathways. We have recently optimized the protein analysis using quantitative MS, allowing precise definition of the dynamic composition of signaling complexes. We now propose to pursue two specific aims: First, to define new components of the key TLR signaling complexes marshaled by MyD88 and TRAF6 using our dimerization technique and, second, to define the function of ABIN-1, a novel TLR signaling component recently discovered by our technique. Preliminary analysis of ABIN-1 deficient mice indicates an inflammatory phenotype with increased lymphoid organ size. PUBLIC HEALTH RELEVANCE: Toll-like receptor family members constitute a major pathogen recognition system, which protects us from infectious diseases via partially characterized signaling pathways. The goal of this project is to reveal the molecular composition of these pathways and to define the function of identified molecules in immune responses. This information will be crucial for our understanding of physiological immune responses, and will also be instrumental for future therapeutic approaches in diseases which are controlled by non-physiological activity of these pathways, i.e. autoimmune diseases and sepsis.
Funding Period: ----------------2009 - ---------------2013-
more information: NIH RePORT

Top Publications

  1. pmc Hematopoietic progenitor cell lines with myeloid and lymphoid potential
    Vanessa Redecke
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, Tennessee, USA
    Nat Methods 10:795-803. 2013
  2. pmc Type I interferon protects against pneumococcal invasive disease by inhibiting bacterial transmigration across the lung
    Kim S LeMessurier
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, Tennessee, United States of America
    PLoS Pathog 9:e1003727. 2013
  3. doi Expanding TRAF function: TRAF3 as a tri-faced immune regulator
    Hans Hacker
    Department of Infectious Diseases, St Jude Childrens Research Hospital, Memphis, Tennessee 38105, USA
    Nat Rev Immunol 11:457-68. 2011
  4. pmc The E3 ubiquitin ligase mind bomb-2 (MIB2) protein controls B-cell CLL/lymphoma 10 (BCL10)-dependent NF-κB activation
    Cinthia C Stempin
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, Tennessee 38105, USA
    J Biol Chem 286:37147-57. 2011
  5. pmc A20-binding inhibitor of NF-κB (ABIN1) controls Toll-like receptor-mediated CCAAT/enhancer-binding protein β activation and protects from inflammatory disease
    Jingran Zhou
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, TN 38105, USA
    Proc Natl Acad Sci U S A 108:E998-1006. 2011
  6. pmc NIK prevents the development of hypereosinophilic syndrome-like disease in mice independent of IKKα activation
    Hans Hacker
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, TN 38105, USA
    J Immunol 188:4602-10. 2012

Scientific Experts

  • Hans Hacker
  • Vanessa Redecke
  • Jingran Zhou
  • Anthony A High
  • Kim S LeMessurier
  • David Finkelstein
  • Liying Chi
  • Ruiqiong Wu
  • Cinthia C Stempin
  • Elaine Tuomanen
  • Vandana Chaturvedi
  • Jerold E Rehg
  • Clive A Slaughter
  • Juan P Giraldo-Vela

Detail Information

Publications6

  1. pmc Hematopoietic progenitor cell lines with myeloid and lymphoid potential
    Vanessa Redecke
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, Tennessee, USA
    Nat Methods 10:795-803. 2013
    ..The simplicity of this system and the unlimited proliferative capacity of Hoxb8-FL cells will enable studies of immune-cell differentiation and function. ..
  2. pmc Type I interferon protects against pneumococcal invasive disease by inhibiting bacterial transmigration across the lung
    Kim S LeMessurier
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, Tennessee, United States of America
    PLoS Pathog 9:e1003727. 2013
    ..e. PAF receptor-mediated transcytosis and tight junction-dependent pericellular migration, ultimately limiting progression from a site-restricted lung infection to invasive, lethal disease. ..
  3. doi Expanding TRAF function: TRAF3 as a tri-faced immune regulator
    Hans Hacker
    Department of Infectious Diseases, St Jude Childrens Research Hospital, Memphis, Tennessee 38105, USA
    Nat Rev Immunol 11:457-68. 2011
    ..In this Review, we discuss our current understanding of the role of TRAF3 in TNFR and TLR signalling pathways, and its role in disease...
  4. pmc The E3 ubiquitin ligase mind bomb-2 (MIB2) protein controls B-cell CLL/lymphoma 10 (BCL10)-dependent NF-κB activation
    Cinthia C Stempin
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, Tennessee 38105, USA
    J Biol Chem 286:37147-57. 2011
    ..Knockdown of MIB2 inhibited BCL10-dependent NF-κB activation. Together, our results identify MIB2 as a novel component of the activated BCL10 signaling complex and a missing link in the BCL10-dependent NF-κB signaling pathway...
  5. pmc A20-binding inhibitor of NF-κB (ABIN1) controls Toll-like receptor-mediated CCAAT/enhancer-binding protein β activation and protects from inflammatory disease
    Jingran Zhou
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, TN 38105, USA
    Proc Natl Acad Sci U S A 108:E998-1006. 2011
    ..Together, our data reveal ABIN1 as an essential anti-inflammatory component of TLR-signaling pathways that controls C/EBPβ activity...
  6. pmc NIK prevents the development of hypereosinophilic syndrome-like disease in mice independent of IKKα activation
    Hans Hacker
    Department of Infectious Diseases, St Jude Children s Research Hospital, Memphis, TN 38105, USA
    J Immunol 188:4602-10. 2012
    ....