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Title page for ETD etd-08032010-133210

Type of Document Dissertation
Author Goettel, Jeremy Allen
URN etd-08032010-133210
Title Kinase suppressor of Ras 1 is a functional protein kinase and protects from experimental colitis in mice by regulating T lymphocyte IFN-γ production
Degree PhD
Department Cell and Developmental Biology
Advisory Committee
Advisor Name Title
Steven K. Hanks Committee Chair
D. Brent Polk Committee Member
Matthew J. Tyska Committee Member
Roy Zent Committee Member
Stephen R. Hann Committee Member
  • MAPK
  • KSR1
  • IFN
  • Colitis
  • IL-10
  • Kinase
Date of Defense 2010-08-02
Availability unrestricted
One Immunological disorders of the gastrointestinal tract is inflammatory bowel disease (IBD). IBD results in recurrent and persistently elevated levels of cytokines, such as tumor necrosis factor (TNF). TNF promotes activation of the mitogen-activated protein kinase (MAPK) pathway that consists of protein kinases Raf/MEK/ERK. Kinase suppressor of Ras-1 (KSR1) binds all three MAPK cascade components Raf, MEK and ERK to facilitate activation of this pathway. While KSR1 possesses a protein kinase domain, mammalian KSR1 contains an arginine in place of an invariant lysine residue required for ATP binding. Thus evidence supporting a catalytic function of KSR1 remains controversial. Using a recombinant protein expression system in bacteria, we demonstrated that KSR1 is a functional protein kinase that undergoes serine autophosphorylation and is phosphorylates recombinant kinase-inactive MEK1 (rMEK1 K97M). In addition, immunoprecipitated FLAG-tagged wild-type KSR1 expressed in KSR1-/- cells also phosphorylated rMEK1 K97M. Finally, KSR1 promoted colon epithelial cell survival in response to TNF that was dependent on a functional KSR1 kinase domain and MEK kinase activity in vitro.

Since TNF is upregulated in human IBD, we studied the role of KSR1 during chronic inflammation by crossing KSR1-/- mice with the interleukin-10 deficient (Il10-/-) mouse model of spontaneous experimental colitis. KSR1-/-Il10-/- mice developed an accelerated severe colitis by 4 weeks of age. We determined that KSR1 expression in hematopoietic lineages mediated significant protection against colitis in Il10-/- mice. Further analysis indicated that naïve CD4+ T cells lacking KSR1 produced higher levels of IFN-γ and had a greater propensity to differentiate along the Th1 axis. Finally, administration of neutralizing antibody against IFN-γ attenuated the disease in KSR1-/-Il10-/- mice.

In summary, we have demonstrated that KSR1 is a functional protein kinase, MEK1 is an in vitro substrate of KSR1, and the catalytic activities of both proteins are required for eliciting cell survival responses downstream of TNF. In addition, KSR1 suppresses colitis in Il10-/- mice by regulating IFN-γ production in CD4+ T cells and promotes effector T cell developmental homeostasis. These findings indicate that KSR1 is an important molecule involved in both epithelial cell survival and immune regulation. Therefore, further investigations utilizing agents that modulate KSR1 expression to suppress inflammation may validate KSR1 as a therapeutic target for patients with IBD.

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