Involvement of Myova and Actin in Insulin Granule Trafficking

Pancreatic beta-cells secrete insulin in response to elevated blood glucose. After synthesis, insulin granules are transported initially by kinesin-I, then transferred to MyoVa within the actin cortex. Granule trafficking models propose that MyoVa tethers granules within the actin cortex, which acts as a secretion barrier. Upon glucose stimulation, actin cytoskeletal reorganization allows granule diffusion or transport by MyoVa to the plasma membrane. To determine how MyoVa and actin contribute to granule trafficking, we tracked eGFP-labeled granules in INS-1 cells (resolution: 23nm/50ms), analyzing their movement by mean square displacement. At rest, granules display 3 modes of motion: paused, diffusive, and directed. Glucose stimulation increases the percentage of granules undergoing MyoVa and/or kinesin-directed motion with little change in the dynamic movements of peripheral actin cytostructures. However, Jasplakinolide treatment, which changes actin's depolymerization rate, inhibits actin cytostructural dynamics and reduces the percentage of granules with directed movement. Therefore, a dynamic actin cytoskeleton is required for efficient granule transport. By introducing Qdot-labeled MyoVa into cells, we will track granules and the steps of associated MyoVa to distinguish between MyoVa's proposed roles as granule transporter or tether.