138 Biochimica et Biophysica Acta, 544 (1978) 138--143 © Elsevier/North-Holland Biomedical Press BBA 28695 SURFACE ANALYSIS AND DEPTH PROFILES OF CALCIUM IN HEPATOMA CELLS DURING PYRUVATE-INDUCED DNA SYNTHESIS L. PICKART, M.M. MILLARD, B. BEIDERMAN and M.M. THALER Department of Pediatrics and The Liver Center, University of California, San Francisco, Calif. 94143, and Western Regional Research Center, U.S. Department of Agriculture, Berkeley, Calif. 94710 (U.S.A.) (Received April 6th, 1978) Summary Induction of DNA synthesis is associated with increased uptake of calcium in cultured cells. Calcium distribution within the plasma membrane and adjacent cytoplasmic layers of hepatoma cells was investigated with X-ray photoelectron spectroscopy and oxygen plasma etching. Cells in minimal growth medium initiate active DNA synthesis 16 h after addition of sodium pyruvate. Cells stimulated with pyruvate and pyruvate-free controls were analysed by X-ray photoelectron spectroscopy--oxygen plasma etching at 0--40/~ (layer I), 0--450 /~ (layer II) and 0--4000 /~ (layer III) from the outer cell surface. Cal- cium concentrations were elevated in induced cells compared with controls: +20% in layer I, +60% in layer II, and +300% in layer III. As the plasma mem- brane is 90--120 A thick, these results indicate that pyruvate-induced DNA synthesis is preceded by an increase in calcium, most marked in the cytoplasm subjacent to the plasma membrane, moderate at its inner surface, and mini- mal at its outer surface. Calcium is known to activate DNA replication in various cultured cell types [1--3]. In addition, enhancement of DNA synthesis by growth factors, lectins and other mitogens appears to be calcium dependent [4,5]. The stimulatory effect of calcium on DNA replication may be mediated at the cell surface, or may be due to direct involvement of calcium in intracellular processes. In order to obtain information which may distinguish between these possibilities, the distribution of calcium at the cell surface and in the cytoplasm was investi- gated in hepatoma cells cultured under basal conditions, and following induc- tion of DNA synthesis by a stimulus other than calcium. The calcium gradient from the external cell surface to the cellular interior was measured with X-ray photoelectron spectroscopy combined with oxygen plasma etching of the cell surface [6,7].