UBC Theses and Dissertations
Regulation of hyaluronan binding by the cell adhesion molecule CD44 in myeloid cells Brown, Kelly L.
CD44 is a cell adhesion molecule that mediates cell-cell and cell-ECM interactions through an association with hyaluronan, the most characterized ligand for CD44. CD44 and HA are ubiquitously expressed, but they do not constitutively interact. At the onset of this work, multiple factors were reported to correlate with the activation and conversion of CD44 to an HA-binding form in various cell types however, minimal data linked specific factors and associated molecular mechanisms to particular physiological stimuli. It has been documented that pro-inflammatory agents induce monocytes to bind to HA. CD44- HA interactions have been implicated in cell adhesion and cell migration events during an inflammatory response and in the pathogenesis of inflammatory disorders. The research described herein aimed to understand the stimuli and corresponding cellular mechanisms that regulate CD44-mediated HA-binding in myeloid cells under inflammatory conditions. The experiments were conducted using a human myeloid progenitor cell line and primary ex vivo human monocytes and murine bone marrow-derived macrophages. Resulting from this investigation, two independent mechanisms that control HA-binding in myeloid cells in response to inflammatory agents were elucidated. One mechanism involves the sulfation of the extracellular domain of CD44. Sulfated CD44 has enhanced reactivity with HA on a cellular and molecular level. Furthermore, a shift in sulfate distribution on CD44 favouring O- and N-linked glycans occurred following cellular activation with inflammatory stimuli, correlating with the induction of HA-binding. A change in sulfate incorporation and distribution on CD44 may encourage conformational changes to CD44 that boost avidity and/or affinity for HA. The second mechanism that was identified to potentially regulate HA-binding in myeloid cells involves an intracellular association between the cytoplasmic domain of CD44 and the cytoskeleton of the cell. A novel CD44-phospho-ERM complex co-localizes to F-actin rich membrane protrusions to promote HA-binding. A mechanism of receptor clustering by the intracellular domain of CD44 likely enhances CD44 avidity for HA. Evidence presented in this thesis demonstrates that the regulatory mechanisms determine different binding affinities for HA that, in turn, confer different adhesive or migratory phenotypes to myeloid cells. In light of recent data that proposes a conflicting role for CD44, that is, in both the onset and resolution of an inflammatory response, this work proposes that the induced interaction between CD44 and HA can support both cell adhesion to, and migration within, the ECM. This provides evidence that CD44 can play a dual and opposing role in an inflammatory response by promoting an initial infiltration of cells to the tissue to generate the inflammatory response, and to promote subsequent cellular adhesion of cells that 'clean-up' and down-regulate the response. Overall, this research offers insight into the complexity of the molecular regulation of HAbinding by the cell adhesion molecule CD44 and implicates CD44-HA interaction in the onset and resolution of inflammation.
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