CD4-positive T cells express ligands for E- and P-selectins, carbohydrate-binding adhesion molecules that are induced on inflamed endothelium. The interaction between selectins and the selectin ligands facilitates the migration of T cells to sites of inflammation. The fucosyltransferase FucT-VII is essential for selectin ligand formation but the signaling and transcriptional pathways regulating FucT-VII induction in T or myeloid cells are poorly understood. We show that constitutively active H-Ras induces FucT-VII transcription in T cells. Retroviral and pharmacological inhibition studies revealed the involvement of the classic Ras-Raf-MEK-ERK signaling cascade and that FucT-VII is specifically induced by the H-Ras, but not the N- or K-Ras isoforms. Pharmacological inhibition studies suggested that phosphoinositide-3 kinase (PI3K) activity is essential for H-Ras-mediated FucT-VII upregulation. However, the H-Ras specificity for FucT-VII induction is not due to differential activation of the Raf or PI3K pathways by the Ras isoforms. Studies employing H-Ras mutants, which interact selectively with certain Ras effectors, and active Raf, suggested that H-Ras-mediated FucT-VII induction requires the concomitant activation of at least three pathways, the Raf, PI3K and a distinct, H-Ras specific, effector signaling cascade. We also investigated the role of the transcription factor T-bet in the expression of glycosyltransferases involved in selectin ligand biosynthesis. T-bet is responsible for the IL-12-mediated, Stat4-independent enhancement of FucT-VII expression in Th1 cells and identified ST3Gal-VI, as a T-bet- and Stat4-dependent gene involved in E-/P-selectin ligand formation. FucT-VII expression is inducible and higly regulated in T cells, but constitutive in myeloid cells. Studies employing adenovirus E1A mutants, which sequester components of the transcriptional machinery, suggested a role for the coactivators of transcription p300/CBP and the chromatin remodeling complexes p400/TRRAP in FucT-VII expression in myeloid cells and indicated distinct modes of regulation in myeloid and T cells. Concluding, H-Ras induces FucT-VII expression in T cells H-Ras by activating the Raf/MAPK and PI3K cascades, as well as a third, H-Ras specific pathway, while IL-12-mediated FucT-VII enhancement in Th1 cells is T-bet-dependent. The transcriptional mechanism of constitutive FucT-VII expression in myeloid cells is distinct from T cells and requires the coactivators p300/CBP and p400/TRRAP.

Last modified

• 05/09/2018

Creator

• Dimitrios G. Zisoulis

DOI

• https://doi.org/10.21985/N2ZH71

Subject

• Integrated Graduate Program in the Life Sciences

Keyword

• Biology

Date created

• 2006-12-11

Resource type

• Dissertation

Rights statement

• In Copyright