Edward D Lipson - US grants

Photoreception and sensory transduction will be investigated in the fungus Phycomyces, in connection with its phototropism and other blue light responses. The long term objective is to understand, at the cellular and molecular levels, the processes by which a stimulus such as light is transduced into a cellular response. The unicellular sporangiophore of Phycomyces serves as a model system for primary receptor cells. The absolute operating range of 10-10:1 in light intensity and the kinetics of light and dark adaptation are similar to those observed in visual photoreceptors; in Phycomyces, the cell is naturally isolated and amenable to genetic approaches. Action spectra for phototropism and the related light-growth response will be measured to characterize the low and high intensity photosystems of the sporangiophore. Experiments with monochromatic and dichromatic continuous irradiation will be applied in a way that allows the determination of components of each photosystem and their interaction with components of the other photosystem. These experiments will also involve single and double pulse stimuli applied to dark adapted sporangiophores of wild type and mutants with altered photoreceptors; the experiments will detect any light-absorbing intermediates. The experiments will be recorded with time-lapse video equipment. Null action spectra for the light-growth response will be measured automatically on the Phycomyces tracking machine; the results will be compared with phototropic balance spectra to determine whether these responses both use the components of the photoreceptor system in the same way. The recently discovered dependence of dark adaptation on low level residual light will be pursued to help establish which photoreceptor mediates this cryptic sensitivity when the sporangiophore is unable to respond directly. System-identification experiments on the light-growth response will be continued on the tracking machine with sum-of-sinusoids test stimuli to measure the dynamic and nonlinear aspects of this response under conditions of wavelength, temperature, and intensity range. These experiments will involve both wild-type and mutant strains, and will be interpreted with analytical models for the kinetics of the photosensory transduction chain.

The objective of this research project is to identify and isolate molecules that serve as blue-light photoreceptors in the fungus Phycomyces. Special attention will be given to phototropism mutants (genes madA to madH) including those (madB and madC) with defective photorecptor(s). Recent spectrophotometric and biochemical analyses have revealed modifications in the photorecptor mutants, notably light-induced absorbance changes (LIAC) with specific alterations in madC mutants. LIAC will be measured in vivo and in vitro and will serve as a spectroscopic assay for the photoreceptor(s). Biochemical characterization and purification of the photoreceptor(s) will employ two-dimensionl gel electrophoresis and fast protein liquid chromatorgraphy (FPLC) in conjunction with spectroscopic and genetic assays. Flavoproteins will receive special consideration as probable candidates for blue light photoreceptors. Electrophoresis and chromatography will also be applied to proteins found to be altered in other behavioral mutants (madA, madB, and madE). Additional mutants will be sought and incorporated in the project. Although blue-light responses are widespread among plants, funji, and other organisms, the photoreceptor molecules remain to be identified. Phycomyces offers special advantages, in view of its unmatched sensitivity and the availability of behavioral mutants. Identification of photoreceptors in Phycomyces will greatly promote the understanding of photosensory transduction in connection with its various light responses (phototropism, light- growth response, photocarotenogenesis, and photodifferentiation). The results on Phycomyces should lead the way to identification of blue light photoreceptors in other organisms.

Support is requested for a preparative ultracentrifuge, liquid scintillation counter and an, autoclave. This equipment is urgently needed for biochemical studies of photoresponses in two eukaryotic microorganisms, the fungus Phycomyces blakesleeanus and the alga Chlamydomonas reinhardtii. In Phycomyces, blue light controls several response, including phototropism, light growth responses, carotene synthesis, and sporangiophore development. The blue-light photoreceptors in Phycomyces and in many other plants and fungi remain to be identified. The availablity of well characterized mutants, provides a strategy for biochemical isolation of photoreceptors and other molecules involved in photosensory transduction processes. In Chlamydomonas, phototaxis and carotene synthesis employ a rhodopsin photoreceptor. This rhodopsin, however, has yet to be purified in sufficient quantity for spectroscopic analysis. The increasing level of biochemistry work in the two biophysics research groups requires that appropriate equipment be available within the physics building, instead of only in the biology building across campus. The ultracentrifuge is required for cell fractionation, in particular membrane fractionation. The liquid scintillation counter will be used for receptor binding studies and for tracing signaling pathways using radioactively labeled probes. The autoclave will be used for sterilization of rowth media and apparatus.

Photoreception and sensory transduction will be investigated for two blue- light responses of the unicellular sporangiophore of the fungus Phycomyces. Both phototropism and the light-growth response occur over an absolute operating range of 10 decades in light intensity from 10-9 to 10 W m-2. This range and the associated adaptation phenomena are similar to those that occur in visual receptor cells. The long term objective of the proposed research is to understand the processes by which a light stimulus is transduced into a cellular response, particularly in organisms with specific blue-light sensitivity associated with flavin chromophores. In Phycomyces, a model organism, genetic and physiological approaches can be effectively combined. Action spectra for phototropism and the light-growth response will be measured in different intensity ranges to characterize the low and high intensity photosystems of the sporangiophore. Phototropism action spectra will be obtained with continuous and pulsed light protocols. Experiments on kinetics of phototropism will be recorded with time-lapse video equipment and analyzed with the aid of an tracking machine using a novel method. The results will be compared with phototropic balance spectra to determine how each of the blue-light responses uses the components of the photoreceptor system. The recently discovered dependence of dark adaptation on dim, subliminal light will be pursued to help establish which photoreceptor mediates this effect, which is sensitive to green and red as well as blue light. System-identification experiments on the light-growth response will be continued on the tracking machine with Gaussian white noise and sum-of-sinusoids test stimuli to measure the dynamic and nonlinear aspects of this response. These experiments will involve both wild-type and mutant strains, and will be interpreted with analytical models for the kinetics of the photosensory transduction chain. A new theory, developed elsewhere, concerning light distribution around a sporangiophore will be tested experimentally by recording the azimuthal light pattern around a sporangiophore illuminated with a horizontal laser beam. This theory is important for interpretation of phototropism action spectra and is fundamental for theories of phototropism. Finally, cellular physiology methods will be applied to sporangiophores and protoplasts. Effector substances and indicator dyes will be introduced by direct uptake, injection, and electroporation; modified blue-light responses and effects will be monitored in preparations from wild type and mutant strains. These approaches are directed towards identifying molecular components and pathways associated with the blue light responses of Phycomyces.

9523481 Lipson This project will create a Metacenter Regional Alliance between the Cornell Theory Center and the Northeast Parallel Architectures Center (NPAC) and the Department of Physics at Syracuse University, to develop interactive multimedia educational modules incorporating advanced scientific simulations. These will be integrated into undergraduate courses, and then into K-12 curricula, to enhance the teaching of general science courses. The project will combine leading-edge technologies in high-performance computing and communications, a high-speed network serving as a testbed for the National Information Infrastructure, and advanced client/server technologies such as the World Wide Web, VRML and Java, to produce and deliver these simulations and educational modules. An important aspect of this project is the conversion of computational science simulations into interactive educational modules. Four simulations are proposed: fluctuating membranes, fluid dynamics, crack propagation and structural failure, and avalanches. The researchers will utilize three different methods to provide these simulations in the classroom: annotated digital video on demand; partially interactive exploration using multiple video streams; and fully interactive simulations on demand. Hypermedia navigation based on the concepts of the World Wide Web will be used to impose a coherent interface to these different simulations, and to provide educational information about the concepts illustrated by the simulations.

We propose to develop, test, and produce a set of educational modules that use computer and network-based resources to teach science topics in an integrated way to non-science majors. These modules will consist of hypertext documents combining text, still images, video, and sound using state-of- the-art technology. Two prototype units covering topics of a successful new two-semester course taught at Syracuse University called Science for the 21st Century have already been tested in the classroom. Advanced information-transfer technologies such as video-on-demand and simulation-on- demand will be implemented to increase active student involvement. New units will be developed to cover other important science topics, such as evolution and imaging science phenomena. The five modules will constitute an introduction to science for undergraduates who are not science majors. Each module will be self-contained and easily expandable; it will be portable since it will be delivered through the Internet or in other formats, such as CD-ROM. There will be three phases of this project: a) design and development of modules, b) implementation, testing, and assessment of modules in our course, and c) dissemination of products and training of teachers. Through this pioneering project, we hope to contribute to the process through which revolutionary information technology connects with improvements in our ability to educate our citizenry.

9554451 Johnson Syracuse University will initiate a 4-week, summer residential, Young Scholars project in interdisciplinary science and mathematics for 50 economically disadvantaged students from New York State entering grade 8. The program will provide high ability and high potential middle school student with early experiences in mathematical problem solving, laboratory science, computer science, and research in a highly charged environment. The summer component combines course work with extensive laboratory and research experience, and includes field trips designed to increase students' interest and awareness of research career paths. Academic year follow-up activities include an extension of activities and research in which the students were involved during the summer.