Contact Inhibition of Locomotion: A Genetic Dissection and Mosaic Model

Contact inhibition of locomotion (CIL) is a phenomenon observed when migrating cells collide and repel each other. Despite having been characterized over 50 years ago, relatively little has been understood about the regulatory mechanisms that govern this essential migratory process. CIL has been demonstrated to be important in development and it has been shown that cancerous malignant cells lose the ability to properly undergo contact inhibition. Previous lab research has demonstrated CIL in vivo to be critical for the migration of embryonic Drosophila hemocytes, which eventually disperse to form a stereotypical evenly spaced pattern. This standardized dispersal lends itself readily to fluorescent screening for regulatory genes involved in CIL and a pilot screen was conducted to test the feasibility of this screening approach. Fluorescently labeled fly lines with various characterized genetic deficiencies were synthesized and migratory hemocytes were imaged using a nuclear marker. Subsequently, the hemocyte migratory pattern was analyzed visually and the spacing examined using a nearest neighbor algorithm. Embryos carrying genetic deficiencies often exhibited severe morphological defects but generally maintained the stereotypical hemocyte migratory pattern, suggesting a degree of independence between hemocyte dispersal and morphological mutations. Due to some amount of overlap between colliding cells it has been difficult to image individual cells during collisions. Therefore, we have also developed an imaging technique yielding single cell resolution of cytoskeletal filaments during collisions. Fly lines that expressed different fluorescent labels for actin and microtubule filaments were synthesized. Fluorescence laser bleaching was then used to eliminate the specific fluorophores belonging to either actin or microtubule filaments. This allows us to create time-lapse mosaic images that capture single cell actin or microtubule filament interactions during collision events. Once genes have been identified as affecting hemocyte patterning, this mosaic imaging assay will be used to understand how these genes affect CIL mechanisms. Further analysis using these imaging techniques will result in the most exhaustive study of CIL to date and will serve to elucidate the molecular and genetic mechanisms involved in CIL. Supported by Minority Health and Health Disparities International Research Training (MHIRT) Program, NIH Grant MD-01485.