Nematodes are the largest population of parasites of the plant and animal kingdoms, serious parasites in humans, and yet few details of their prodigious reproduction are known. The long-range goal of this work is to understand the morphogenesis, locomotion, chemotaxis and fertilization by the nematode spermatozoan. Nematode sperm are amoeboid rather than flagellated, and crawl rather than swim. The locomotive machine is its permanent extended pseudopod; the continuous flow of membrane under it pulls the cell body long. Ascaris suum sperm are used here as a model for amoeboid sperm motility. In the presence of an extract from the male reproductive tract, inactive sperm cells form pseudopods in vitro (Foor and McMahon, 1973), eventually become fully motile and crawl rapidly under proper conditions. In living cells, membrane specializations (villipodia) continuously form at the tip of the pseudopod, migrate rapidly rearward and disappear at the junction between pseudopod and cell body (the """"""""CPJ""""""""). Using hog-voltage EM (HVEM), an interconnected cytoskeleton of fiber complexes was found within the pseudopod and villipodia of motile Ascaris spermatozoa. It is visible in living, creeping sper and, surprisingly, the entire cytoskeleton moves rearward in unison with the rapid membrane movement. The processes driving this internal movement and the membrane flow itself are not known. It is the specific purpose of this grant to study the distinctive properties of the pseudopod membrane and the source of its continuous evolution. Recently, an unusually dense accumulation of tubular and round membrane vesicles ws discovered at the CPJ. It is thought to be part of a postulated membrane-recycling system that maintains continous membrane formation at the tip and intermalization at the CPJ. Experiments are proposed to: (1) examine the anatomical relationship of the pseudopod plasma membrane to the cell body PM and the CPJ vesicles using HVEM and low-voltage SEM; (2) isolate plasms membranes and CPJ vesicles from inactive and activated cells for SDs-gel electrophoresis and specificafinity lavelling to characterize membrane proteins; (3) make a pane; of monoclonal antibodies which recognize specific membrane fractions; (4) labels in vivo activating sperm to determine the membrane domains during activation and crawling. These antibodies will be useful in later studies for isolating plasma membrane proteins associated with adhesion and, possibly, linkage to the cytoskeleton.
