THE RON RECEPTOR TYROSINE KINASE, A NEGATIVE REGULATOR OF INTERFERON-GAMMA AND ENDOTOXIN INDUCED MACROPHAGE ACTIVATION
Open Access
- Author:
- Ray, Manujendra
- Graduate Program:
- Integrative Biosciences
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- May 15, 2008
- Committee Members:
- Pamela Hankey Giblin, Committee Chair/Co-Chair
Avery August, Committee Member
Robert Paulson, Committee Member
Margherita Teresa Anna Cantorna, Committee Member
Aimin Liu, Committee Member - Keywords:
- Receptor Tyrosine Kinase
RON
Lipopolysaccharide
Interferon Gamma
Endotoxic Shock
Inflammation - Abstract:
- Receptor tyrosine kinases (RTKs) are emerging as a class of key regulators of innate immune responses. The RON RTK is expressed on tissue-resident macrophages, and inhibits classical macrophage activation while promoting hallmarks of alternative activation, thus regulating the critical balance between the inflammatory and wound-healing properties of activated macrophages. Here we demonstrate that, in response to in vivo lipopolysaccharide challenge, RON-/- mice harbor significantly increased systemic levels of IFN-and IL-12, and increased levels of IL-12p40 transcript in their spleen. This elevation of IFN-can be attributed to splenic NK cells responding to the elevated levels of IL-12. Analysis of RON/IFN-γ receptor double knockout mice indicates that the enhanced susceptibility of RON-/- mice to endotoxin challenge is dependent on IFN-receptor mediated signals. In vitro studies demonstrate that stimulation of primary peritoneal macrophages with macrophage stimulating protein (MSP), the ligand for RON, inhibits IFN--induced STAT1 phosphorylation and CIITA expression, resulting in reduced surface levels of MHC Class II, a classic IFN- responsive gene. Taken together, these results indicate that the RON receptor regulates both the production of, and response to, IFN- resulting in enhanced susceptibility to endotoxin challenge. In vivo following LPS challenge, RON-/- mice harbor elevated levels of IL-12. We also show here that signaling through MSP/RON inhibits LPS induced IL-12p40 expression at least in part by inhibiting various aspects of the TLR-4 and TLR-4 induced IFN-β signaling pathways. Our results demonstrate that signaling through RON not only retards LPS induced IkB-α degradation but also inhibits the phosphorylation of p65 serine 536. While signaling through MSP/RON has no effect on LPS induced IRF-3 activation, it does however inhibit LPS/IFN-β induced STAT-1 tyrosine phosphorylation. In addition as we find that LPS induces RON expression, this suggests that RON functions in a negative feed back loop to inhibit LPS signaling. Finally, at a receptor proximal level we show that the second docking site tyrosine of RON and the GRB-2 adaptor protein are required for the MSP/RON mediated inhibition of LPS induced IL-12p40. The stimulation of macrophages with MSP results in the induction of SOCS1 and SOCS3. These data are highly correlative with our findings that signaling via MSP/RON induces ERK and STAT-3 activation, both of which have been shown to be important for SOCS expression. The manner in which RON inhibits LPS and IFN-γ induced signals is very similar to the manner in which the SOCS proteins do so. It therefore seems likely that signaling through RON indirectly inhibits IFN-γ and LPS induced signals via SOCS1 and SOCS3. Taken together, work done in this dissertation provides an insight into the mechanism by which RON regulates LPS induced, and IFN-γ potentiated macrophage activation and the systemic effects of this regulation.