A great deal is known about the atomic structures and biophysical mechanisms of motor proteins. However, much less is known about how motor proteins are regulated by intracellular signals and how such signals impinge upon and coordinately turn on and off kinesin and dynein, two opposite polarity microtubule motor proteins. We are using a synthetic biology approach for tackling these problems by creating artificial cargos that are transported by motor proteins inside of cells and, like natural organelles, are responsive to intracellular signals. Using light microscopy and structural biology approaches, we aim to develop a mechanistic understanding of how motor proteins are regulated.