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Molecular interactions and signaling mediated by the cell adhesion molecule CD44

University of British Columbia

CD44 mediates cell adhesion and cell migration in leukocytes. Most of these functions involve the interaction of cell surface CD44 with one of its most characterized ligands, hyaluronan or HA, a component of the extracellular matrix. The ability of CD44 to bind HA is strictly regulated in leukocytes such that not all CD44-expressing cells will bind HA. All three domains of CD44, the extracellular domain, the transmembrane domain, and the cytoplasmic domain, regulate HA binding. The extracellular domain of CD44 has been studied extensively for its regulation on HA binding, however, the mechanisms by which the transmembrane and the cytoplasmic domains of CD44 influence HA binding are poorly understood. In this study, therefore, I have focused on the roles for the transmembrane and cytoplasmic domains of CD44 in regulating HA binding and signaling in leukocytes. Using chimeric CD44 molecules, the transmembrane domain of CD44 was shown to be responsible for the non-covalent association of CD44 in the plasma membrane of T cells. This self-association of CD44 was found to facilitate HA binding. In addition, CD44 was also observed to associate with the cytoskeletal components, ezrin/radixin/moesin (ERM), with Srcfamily kinases, Lck and Fyn, and with an adapter protein, Grb-2. CD44 interacted with ERM proteins in T cells and myeloid cells. In vitro, this interaction required a low ionic strength and the presence of EDTA, as well as the integrity of actin filaments. The interactions of CD44 with Src-family kinases and Grb-2 took place in T cells. CD44 interacted with the Src-family kinase Lck in the membrane lipid microdomains, which are important for receptor signal transduction. In BW5147 T cells, this CD44-Lck interaction, as well as the distribution of Lck in lipid microdomains, is negatively regulated by CD45, a leukocyte specific tyrosine phosphatase. More strikingly, in the absence of CD45, BW5147 T cells underwent cell spreading concomitant with the tyrosine phosphorylation of Pyk2 and FAK upon stimulation by an anti-CD44 antibody. Inhibition of the activity of Src-family kinases abolished CD44- mediated cell spreading as well as the tyrosine phosphorylation of Pyk2 and FAK. Therefore, signals via cell surface CD44 may induce the tyrosine phosphorylation of Pyk2 or FAK through the interaction with Src-family kinases, which then triggers downstream cellular events, leading to cell spreading. CD45 acts as a negative regulator of this process by inhibiting the molecular interaction of CD44 and Src-family kinases. In summary, this study identified a regulatory mechanism for HA binding by the transmembrane domain of CD44, as well as the molecular interactions and the signaling events mediated by CD44 in leukocytes.

Thesis/Dissertation

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2009-10-08

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http://hdl.handle.net/2429/13756

Microbiology and Immunology

University of British Columbia

UBCV

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