Sulfur has a particularly rich biochemistry and fills a number of important roles in biology. In situ information on sulfur biochemistry is generally difficult to obtain because of a lack of biophysical techniques that have sufficient sensitivity to molecular form. We have recently reported that sulfur K-edge X-ray absorption spectroscopy can be used as a direct probe of the sulfur biochemistry of living mammalian cells [Gnida, M., et al. (2007) Biochemistry 46, 14735-14741]. Here we report an extension of this work and develop sulfur K-edge X-ray fluorescence spectroscopic imaging as an in vivo probe of sulfur metabolism in living cells. For this work, we have chosen onion (Allium cepa) as a tractable model system with well-developed sulfur biochemistry and present evidence of the localization of a number of different chemical forms. X-ray absorption spectroscopy of onion sections showed increased levels of lachrymatory factor (LF) and thiosulfinate and decreased levels of sulfoxide (LF precursor) following cell breakage. In intact cells, X-ray fluorescence spectroscopic imaging showed elevated levels of sulfoxides in the cytosol and elevated levels of reduced sulfur in the central transport vessels and bundle sheath cells.

Sulfur is essential for life, with important roles in biological structure and function. However, because of a lack of suitable biophysical techniques, in situ information about sulfur biochemistry is generally difficult to obtain. Here, we present an in situ sulfur X-ray absorption spectroscopy (S-XAS) study of living cell cultures of the mammalian renal epithelial MDCK cell line. A great deal of information is retrieved from a characteristic sulfonate feature in the X-ray absorption spectrum of the cell cultures, which can be related to the amino acid taurine. We followed the time and dose dependence of uptake of taurine into MDCK cell monolayers. The corresponding uptake curves showed a typical saturation behavior with considerable levels of taurine accumulation inside the cells (as much as 40% of total cellular sulfur). We also investigated the polarity of uptake of taurine into MDCK cells, and our results confirmed that uptake in situ is predominantly a function of the basolateral cell surface.