Following Actin Fibers in 3D During Cell Migration in Collagen Matrices

Abstract

Actin polymerization is a major mechanism for the production of the force necessary for cell migration in 2D. The polymerization of actin and its retrograde motion at the leading edge of cell moving in 2D has been studied in great detail as well as the interaction of actin with focal adhesions. When cells grow in 3D collagen matrices, the extending lamellipodial protrusion is more difficult to visualize and it is likely not relavent for the movement of the cell over large distances. We use the modulation tracking 3D method to accurately image the cell protrusion. This method is capable of producing detailed images of 3D structures at the nanoscale and at the same time measure diffusion and aggregation of molecules in these structures. In 3D, cells produce very long protrusions that presumably grab on the surrounding collagen fibers to propel the rest of the cell body in a specific direction. What is unclear is if the cell is capable of forming stable adhesions with the collagen matrix and how the force is generated. Using the modulation tracking imaging method we can follow the changes in shape of the cellular protrusion and also image separately various proteins, including actin in the cytoplasm and in the membrane. In the thin long protrusion we observe both fast diffusing actin molecules and also relatively immobile species, presumably part of the actin cytoskeleton. We are developing a method to directly measure the movement of the entire actin bundle inside the very thin cells protrusions. The method is conceptually similar to speckle imaging; however, it works in 3D.