Nuclear envelopes were isolated from purified rat and calf liver nuclei via different methods. Basic characterization of nuclear envelopes demonstrated phospholipid distributions similar to those found in microsomes, but enzymatic compositions and fatty acid moieties of phosphatidylserine differed from those in microsomes. A substantial Mg2+-dependent NTPase was found in nuclear envelopes from both sources. The activity was linear with protein concentration and showed a sharp pH dependency with maximal activity near pH 7.5. Arrhenius analysis of the activity in rat liver preparations disclosed an activation energy of 13.8 kcal/mol, and Lineweaver-Burk plots showed a Km of 1.8 mM ATP. Under similar conditions, calf liver preparations showed an activation energy of 13.3 kcal/mol and a Km of 1.9 mM ATP; with Mg2+ added in 5 mM excess (over nucleotide concentration) they yielded linear Eadie plots. The NTPase activity in nuclear envelopes from both sources showed a broad substrate specificity and induced declines in the energy charge of various nucleotide additives that paralleled stimulation of RNA transport in vitro by these additives. Nuclear envelopes from both sources were able to hydrolyze the high-energy phosphate bonds of diphosphate nucleotides. The ability to utilize ADP was not dependent on coupled oxidative phosphorylation or on electron transport; rather, it apparently proceeds via a myokinase-like activity that furnishes ATP. A number of agents that modify RNA transport in vitro similarly modified the NTPase activity. For instance, cAMP increases RNA transport and the NTPase activity, and further investigation showed that cAMP increased the Km of the NTPase activity only slightly and the Vmax by 65%. Further studies in vivo following CC14 or thioacetamide treatment of rats demonstrated a parallelism between alterations in RNA transport in vivo and nuclear envelope NTPase activity. Histochemical studies demonstrated that the NTPase activity was distributed along the nuclear envelope and was not localized to nuclear pores under the conditions employed. Supporting this result, we found that thioacetamide-induced nuclear swelling produces changes in nuclear envelope surface area which parallel increases in the nuclear envelope NTPase activity produced by this treatment. The reciprocity between the NTPase activity in nuclear envelopes and RNA transport, with regard to substrate behavior and to effects of activators and inhibitors and perturbations induced by in vivo treatments, suggests that this activity participates in RNA transport.
All Science Journal Classification (ASJC) codes