The major research emphasis in the Membrane Biology Laboratory from 1969 to1995 was the study of membrane changes and membrane traffic in cells. For our studies we chose to investigate the free living protozoan, Paramecium. We focused particularly on the phagosomal and the endosomal systems. We were also interested in how the membrane's biosynthetic pathway was involved in these processes. We were able to follow in detail the formation of phagosomes, their transformation first into phagoacidosomes and then into phagolysosomes and finally the fate and recycling of the spent phagolysosome membrane at the cytoproct and its return to the cytopharynx for reuse. To study these phenomena we relied extensively on electron microscopy and monoclonal antibody localization of antigens. We used transmission and scanning electron microscopy, freeze fracture, quick-freeze deep-etch techniques and immunogold labeling on cryo-thinsections. We also made extensive use of fluorescence microscopy and video-enhanced microscopy. Collaborations with Dr. Agnes K. Fok made it possible for our lab to be able to isolate and characterize cytoplasmic dynein, a molecular motor, from Paramecium and in isolating axonemal dyneins and to show that these complex macromolecules are preassembled in the cytosol before they are moved into the cilium to be attached to the axoneme (Fok et al., 1994). Recently, we have focused more on the possible causes behind membrane tubulation and the potential importance of membrane tubulation for cell function.
Areas of interest for future work are in isolating the various pools of vesicles and in improving the isolation techniques of digestive vacuoles at given ages. Using these isolated vesicular pools we can begin to investigate, in cell free systems, the very specific macromolecular mechanisms of docking and fusion of membranes in Paramecium. We can also use these pools of membranes for electrophysiological and patch-clamp studies of the ion channels and electrogenic pumps found in the membranes of this cell's organelles. Results obtained from these studies will provide a better understanding of the role of membranes in the phagosome-endosome system of cells. (Supported in the past by the National Science Foundation, USA)
Figure 1 in Allen and Fok, 2000.