I. Chitin synthetase and sensory tranduction processes in Phycomyces. II. The avoidance response, the house growth response and the rheotropic response of Phycomyces

Part I: Chitin Synthetase and the Sensor Transduction Processes in Phycomyces: The response of Phycomyces sporangiophores to various stimuli shows up as changes in the elongation rate of the cell wall, a structure mainly composed of chitin fibrils. The enzyme chitin synthetase was chosen as the subject of this study for the possible role that regulation of its activity might play in the behavioral output. Properties of the enzyme: A simple assay for chitin synthetase has been developed for the flycomyces system. Enzyme prepared from Phycomyces was found to catalyze the synthesis of chitin from UDP-N-acetyl-D-glucosamine. The ion requirements, temperature dependence, buffer and pH dependence, and kinetics for this enzyme were investigated. An antibiotic Polyoxin D was found to be a competitive inhibitor of this enzyme. Cellular localization: To localize the enzyme in the cell: (1) Phycomyces homogenates were fractioned through a series of differential aril isopycnic centrifugations. Each fraction was assayed for its specific activities of chitin synthetase and some membrane marker enzymes. The results suggest that chitin synthetase is a plasma membrane bound enzyme. (2) Autoradiography studies of the sporangiophone showed that this enzyme is located mainly in the growing zone of this structure. Regulations: In vivo, Phycomyces shows a positive growth response to blue light. It is demonstrated that blue light can increase the chitin synthetase activity in vitro This finding supports the idea that regulation of chitin synthetase activity plays a central role in the responses of Phycomyces sensory stimuli. Finally, the possibility of using the chitin synthetase assay as an in vitro photoresponse system in the dissection of the sensory transduction processes of Phycomces is discussed. Part II: The Avoidance Response, the House Growth Response, and the Rheotropic Response of Phycomyces: If an object is placed about 1 mm from the growing zone of a Phycomces sporangiophore growing in air, in about 2 minutes the sporangiophore starts to bend away at a rate of about 2°/minute for as long as half an hour or more. This is called the avoidance response of Phycomyces. The purpose of this study is to find out how a sporangiophore detects a nearby object and avoids it. Electric field, electromagnetic radiation (UV, visible and IR), temperature, humidity and pressure are all excluded as the avoidance eliciting signal. The avoidance response is found to be dependent on the size and the distance of the nearby objects and independent of the compositions and surface properties of the objects. An air current parallel to the barrier and the sporangiophore can eliminate the avoidance response. In conjunction with this key observation, the Phycomyces responses to wind (rheotropic response and wind growth response) and to enclosure (house growth response) were characterized. The key parameter in all these responses seems to be the air movement in the vicinity of the sporangiophore growing zone. All observations are compatible with the assertion that faster growth is associated with slower wind velocity. To specify in which way air movement can become a signal directly received by the sensor, we propose that the avoidance is mediated by a volatile growth effector emitted somewhere along the sporangiophore and sensed by the growing zone (the chemical self guidance hypothesis). Various possible alternative forms of the hypothesis were tested experimentally. The only remaining viable one is that the sporangiophore emits volatile growth promoting molecules continuously. The majority of the molecules are readsorbed by the growing zone before they diffuse away, and the barriers modify the distribution of the molecules by altering the ambient wind pattern. Future tests are discussed and a quantitative formulation of the model is presented in Appendices B and C