The applicants have shown that toxoplasma actively invade host cells to establish intracellular residence, and that certain specialized organelles, the conoid and rhoptries, function coordinately during invasion. The conoid is extended, the rhoptries secrete their lytic contents, and the twisting motility of the parasites propels them into the cells. The applicants now plan to study the effects of antibody on these processes to analyze precisely how it functions in host defense to prevent invasion, i.e. which of these several mechanisms operating in cellular penetration are inhibited by antibody. In addition, the applicants plan to examine in greater detail the basic mechanisms involved in motility, conoid extension, and rhoptry secretion. The presence in toxoplasma of a dynein-like ATPase suggests that microtubles may have an active role in cell movement. Consequently, they have designed experiments that reactivate such an ATPase in detergent-extracted toxoplasma to evaluate any resulting change in motility and in the cytoskeleton. In additional experiments, they plan to identify microfilament networks by simultaneous extraction and fixation, followed by negative staining. To determine whether there is an actinmyosin contractile system involved in the movements of toxoplasma, they will attempt to localize actin in glycerinateds toxoplasma with myosin subfragment-1. Inhibitors of microtubules (colchicine and vinblastine) and an inhibitor of microfilaments (cytochalasin D) will be tested for their effects on the motility of toxoplasmas and on their cytoskeleton. The action of the calcium inonphore A23187 on rhoptry secretion will be evaluated to determine whether calcium is requested for secretion, as in many other secretory cells. Dibucaine, which induces synchronous secretion of mucocysts in Tetrahymena, will also be used to stimulate rhoptry secretion in toxoplasma. The disappearance and reappearance of intramembranous """"""""fusion rosettes"""""""" will be monitored by freeze-fracture techniques for electron microscopy to analyze the depletion and renewal of secretory organelles following treatment with ionophore and dibucaine.
| Skorich, D N; Chiappino, M L; Nichols, B A (1988) Invasion of the guinea pig conjunctiva by Toxoplasma gondii. Invest Ophthalmol Vis Sci 29:1871-80 |
| Nichols, B A; Chiappino, M L (1987) Cytoskeleton of Toxoplasma gondii. J Protozool 34:217-26 |
