The role of interactions between 4N1K and CD47 on integrin activation and integrin-mediated cell adhesion

• Main Author: Leclair, Pascal
• Language: English
• Published: University of British Columbia 2014
• Online Access: http://hdl.handle.net/2429/47040

Integrins are heterodimeric, cell-surface receptors that play a role in adhesion and chemoresistance. Integrins can be found in one of two states: an inactivated state where integrin-mediated adhesion is not supported, or in an activated state where integrin-mediated adhesion is possible. CD47 is a cell surface receptor that is said to regulate integrin functions by interacting with integrins and regulating their activation state. A thrombospondin C-terminal motif, RFYVVMWK, has been found to be responsible for cell adhesion and was subsequently shown to be specific for CD47. The 10-mer peptide derived from this sequence, 4N1K, has since been used as the prototypic ligand for CD47 and this interaction has been shown to decrease integrin-mediated adhesion and induce cell aggregation and apoptosis. However, the role of the 4N1K/CD47/integrin axis in chemoresistance has not been investigated. My thesis investigated the consequences of CD47 ligation by 4N1K on integrin-mediated functions. I found that the 4N1K peptide induces binding of a variety of antibodies, including non-specific control antibodies, to the surface of cells. In addition, cells that were deficient in CD47 expression were able to bind substrate-immobilized 4N1K as efficiently as their CD47-expressing parental cells. 4N1K was also found to block cell adhesion and induce cell aggregation in a manner that was independent on CD47 expression. These results suggest that 4N1K may produce artifacts in assays that use antibodies as reporters of integrin activation due to its hyper-adhesive nature, resulting in 4N1K binding to a variety of Ig-containing proteins, such as antibodies and cell-surface proteins. In addition, non-specific binding of 4N1K on cell surfaces and homotypic 4N1K interactions appear to be responsible for many of the observed phenotypes on cell adhesion and aggregation, and may explain the reported CD47-independent effects of 4N1K. As such, I propose that the 4N1K peptide not be used as a ligand to assess the role of thrombospondin/ CD47 interactions on cell functions, since the non-specific adherent nature of 4N1K could lead to erroneous interpretations of experimental data. === Medicine, Faculty of === Graduate