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Very low concentrations of histones stimu-guanidine, 2,4-dinitrophenol, and oligomycin). The late the release of K+ from mitochondria by an energy-phenomenon is also inhibited by chlorpromazine, dependent process. Either electron transport or adeno-atractyloside, and the divalent cations, Mg2+, Sr2+, sine triphosphate can serve as the energy source. The and Mn2+. Inorganic phosphate, in concentrations histone-induced K+ efflux is blocked by inhibitors of significantly lower than that required for oxidative electron transport (antimycin A, cyanide, and amytal) phosphorylation, is necessary for optimal histoneor by energy-transfer inhibitors or uncouplers (octyl-induced Kf efflux.

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READ, R. S. B., AND MAURXTZEN, C. M. Isolation and preservation of cell nuclei for studies on RNA polymerase activity. Can. J. Biochem. 48, 559-565 (1970).The suitability of saponin for the isolation and of glycerol for the preservation of mammalian cell nuclei has been investigated. The nonionic detergent saponin was found to be a useful e l l lytic agent in a procedure for the isolation of nuclei from several mammalian cell types. The RNA polymerase activity of rat liver nuclei was not affected by treatment with saponin or with some other nonionic detergents that were tested. Low concentrations of ionic agents also did no't affect the activity of the enzyme though higher concentrations caused lysis of the nuclei. The preservation of structure and enzyme activity in the isolated nuclei was achieved by storage at low temperature in a medium containing 70 "/, glycerol together with a suitable concentration of a divaknt metal and phosphate buffer.The DNA-dependent synthesis of RNA is an important step in the genetic expression of cells, and the factors which control the activity of this enzyme are of considerable interest. Thein vitro study of RNA polymerase activity (EC 2.7.7.6) in mammalian cell nuclei requires the prior isolation and purification of these organelles. However, little is known about the effects of different isolation procedures on the RWA polymerase activity of cell nuclei, nor have conditions for their storage with the preservation of enzyme activity been explored.A large number of methods for the isolation of nuclei have been described. Many of these employ mechanical shear for the rupture of wlls, while others rely on detergents for the lysis of cells and the release of nuclei (1-5). It has been reported (6) that the presence of detergents can affect the assay of RWA polymerase in nuclei. It is important, therefore, to determine if the use of detergents for the isolation of nuclei does influence their RNA polymerase activity. Saponin, a plant glycoside extract with detergent properties, is well known as a hemolytic agent, but has not yet been employed for the isolation of nuclei for RNA polymerase studies. The usefulness of this detergent was therefore investigated and compared with that of some other detergents.Also, in this investigation, a method for the storage of isolated nuclei has been sought. The WRCC Postdoctoral Fellow, 1968-1969 value of such a method lies in the saving of time involved in the preparation of nuclei and in the avoidance of possible variations between batches of nuclei. It has been found (7) that the presence of glycerol in the suspending medium enables nuclei to be stored at temperatures below freezing without loss of their structure. Glycerol solutions have also been used for the storage of RWA polymerase from bacterial sources (8). A medium containing 38% glycerol has been used for the storage of chick oviduct nuclei at -20 "C (9). RNA polymerase activity was retained by these nuclei but no data concerning the degree of preservation of the enzyme activity were given...