Noninvasive imaging tools for the visualization of cellular communication

Cell-cell contacts are crucial in a myriad of biological settings including cancer progression,embryonic development, and immune system function. Understanding such processes requiresexamining cellular communication in its native context. Optical imaging tools are ideally suitedfor this purpose.The most commonly employed optical imaging technology is fluorescence. Whilefluorescent tools are useful at the microscopic level, issues of autofluorescence and therequirement for incident light limit their utility at larger length scales. This leaves a gap in ourability to “see” at the macroscopic level.A complementary strategy, bioluminescence, circumvents these issues allowing chemicallygenerated light from a luciferase enzyme to illuminate cellular movement. Bioluminescence hasbeen employed to monitor both intracellular and extracelluar events, from gene expression toprotein secretion.Despite its versatility, bioluminescence has not easily transitioned to monitoring direct cellcontacts. This is due, in part, to a lack of tools that can illuminate celllular interactions at themicroscopic level. Moreover, there are a limited number of luciferase probes useful for cellxiiitracking. My work was aimed at filling this gap by developing new bioluminescent tools forstudying cell populations.To enable visualization of cell contacts, I employed a split-reporter strategy wherein theluciferase halves were tethered to separate cells. This method enabled light emission to be linkedwith the microscopic process of cell contacts. It could be used to monitor immune and cancercell interactions during disease progression.To develop new cell tracking tools, I exploited an enzymatic chemical attachment method toappend bioluminescent enzymes to small molecules and proteins. Furthermore, this method wasused to directly label cell surfaces with luciferases. Such tools will facilitate the monitoring cellsrecalcitrant to genetic manipulation, including stem cells and primary cells, as well as trackingendogenous cell populations, such as metastatic tumor cells.My work will enable the study of cellular contacts as well as non-genetically encodable cellpopulations in vivo and noninvasively. Future work from our lab will use these methodologies tofurther diversify bioluminescent imaging techniques and use the tools I have developed tomonitor stem cell and cancer cell migration in mouse models