Regulation of a self-generated attractant gradient by the extracellular matrix - The graded distribution of attractant cues guides migrating cells to their ultimate destination during embryogenesis, in immune system response, and when cancer cells metastasize. Despite the broad importance of cell migration, it remains poorly understood how stable extracellular attraction gradients are formed and maintained in vivo. The migration of the posterior lateral line primordium in zebrafish embryos is an excellent model to address this question because the primordium can be imaged at high spatial and temporal resolution and manipulated genetically with spatial and temporal control. The primordium expresses the Cxcl12a (Sdf1a) chemokine receptor Cxcr4b and migrates along a stripe of uniform Cxcl12a expressing cells. As the primordium migrates, it generates a linear signaling gradient across itself. The formation of a linear gradient requires the presence of a local source and a local sink. The Cxcl12 clearance receptor, Ackr3b (Cxcr7b), is expressed only in the back of the primordium and serves as a local sink. However, the Cxcl12a expression stripe does not constitute a local source. Therefore, it is unclear how the primordium generates a linear Cxcl12a gradient. One possibility is that Cxcl12a is normally sequestered by the extracellular matrix and that the migrating primordium employs a mechanism to locally liberate Cxcl12a to create a local attractant source. Indeed, my preliminary investigation suggests that the extracellular matrix negatively regulates Cxcl12a availability and that Cxcl12a binding to proteoglycan GAG chains is required for proper migration. In this project, I will determine whether Cxcl12a binding to GAGs is required for the formation of a linear Cxcl12a gradient (Aim 1) and whether heparan sulfate proteoglycans are required to tether Cxcl12a along the migratory route (Aim 2). Finally, I will investigate the mechanism by which the primordium releases bound Cxcl12a to generate a local source (Aim 3). To complete this project and prepare myself for a career as an independent investigator, I have created a training plan to develop expertise in genetics and quantitative imaging. My choice of mentor, collaborators, laboratory and institution ensures I have the resources and support necessary to perform the work described here and to receive meaningful feedback from developmental geneticists and microscopy experts throughout the project.
