Scintillation and autoradiographic studies on 63 nickel uptake in Pseudomonas tabaci

Abstract

Scintillation studies on the uptake of 63Ni2+ by Pseudomonas tabaci demonstrate an incorporation of approximately 2.5 nmol per 10(10) bacterial cells, in medium containing 12 nmol (8.3 microCi) per ml. Over 80% of the incorporated Ni2+ is lost from the cells during washing, fixation and dehydration with ethanol. The remaining insoluble (bound) 63Ni2+ has the highest level in cells fixed in acetic acid/ethanol (0.4 nmol/10(10) cells), with smarter amounts in paraformaldehyde- and glutaraldehyde-fixed cells. The radioactive level in aldehyde-fixed cells represents a total Ni2+ uptake of about 10(−18) g or 10(4) atoms per cell. Light- and electron-microscope autoradiography corroborated the scintillation studies in demonstrating a higher retention of label by cells fixed in acetic acid/ethanol, possibly reflecting a higher retention of medium Mr proteins with this type of fixation. High-resolution electron-microscope autoradiography involving gold latensification with physical development demonstrated a clear localization of silver grains to the central nucleoid region (seen most clearly over the discrete nucleoid of aldehyde-fixed cells) and within this to the chromatin (seen most clearly over the condensed chromatin of acetic acid/ethanol-fixed cells). It is suggested that the incorporated 63Ni2+ labels mainly central, genetically inactive DNA, while peripheral, actively transcribing DNA has little associated radioactivity. The pattern of cation association seen in this bacterium shows a number of close similarities to the situation seen in dinoflagellate cells.