How We Study Protein Transport
Mary L. Preuss, Peggy Weidman and Erik Nielsen
For the greater part of the last century, research in the field of protein transport was synonymous with microscopy. Before the end of the century, this view was dramatically changed by the emergence of innovative genetic, molecular and biochemical approaches that revolutionized and invigorated the field. Far from being displaced as an essential tool, microscopy techniques have also evolved. What was once largely a science of “dead cells” has been transformed into a window on the inner workings of living cells. The objective of this chapter is to provide an overview of the major approaches that are employed in the analysis of protein transport within the membrane trafficking system of eukaryotic cells. In particular, we discuss the identification of several of the common model cargo proteins for studying both secretory and endocytic membrane trafficking in both mammalian and yeast systems. We then discuss the development of both in vivo and in vitro techniques to study the transport of these model cargo proteins within cells, and explain some of the common principles involved in these assays. Finally, we discuss some of the recent advances in imaging techniques and technology that have driven the recent “renaissance” in the use of microscopic techniques in the investigation of membrane trafficking events in living cells.