SPECTROSCOPIC SPECIATION OF SULFUR IN LIVING MAMMALIAN CELLS: HIV & APOPTOSIS

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Sulfur is a vitally important element, with roles in structure, catalysis and metabolism in all organisms. We propose to use sulfur K-edge X-ray absorption near-edge spectroscopy to monitor and to improve our understanding of sulfur metabolism in mammalian cells, as a function of cellular development and in response to various stimuli. In particular, changes in the overall redox environment of the cell, and in organelles such as mitochondria, are thought to be important in controlling different developmental stages of the cell, from proliferation, differentiation, apoptosis and finally to necrosis. Research in this area is in its infancy, and direct information on the sulfur metabolome of the cell would be very valuable. Apoptosis, the process by which cells actively commit suicide, is very important in numerous human diseases, including HIV, where induction of apoptosis is the primary cause of T-cell death, and in cancer, where apoptosis is abnormally reduced. Apoptosis is thought to be initiated by redox changes within cells, although direct evidence for this is lacking. Such changes should be measurable by the experiments that we propose, and vital information on this important biological mechanism can anticipated. We propose to use XAS to study cultures of living mammalian cells - the MDCK immortalized cell-line, which are polarized epithelial cells. We have already developed the techniques and equipment for handling the cell cultures and recording their XAS spectra. We have also established the use of sulfur K-edge X-ray absorption near-edge spectroscopy to quantitatively speciate the sulfur in living systems. Now we plan to use these tools to study sulfur biochemistry as a function of cellular developmental status including apoptosis and during mechanical stimulation. Our experiments should provide valuable insights into the role of sulfur in these processes, and particularly in the case of apoptosis, could have important health-related ramifications.