Apparent induction by phenobarbital of uridine diphosphate glucuronyltransferase activity in nuclear envelopes of embryonic-chick liver.

The similarity in enzyme profile between microsomal preparations and nuclear envelopes has been shown for a range of enzymes in adult liver of various species (Franke, 1974; Kasper, 1974). The degree to which the enzymes in these two membrane systems can be induced is, however, variable. In rat liver, phenobarbital produced a two to three times increase in NADPH-cytochrome c reductase (EC 1.6.2.4), aryl hydroxylase (EC 1.14.14.1) and N-demethylase activities in the microsomal fraction compared with controls, but little change in nuclear envelope (Kasper, 1971), whereas in rabbit liver the induction of NADPH-cytochrome c reductase, benzopyrene hydroxylase (EC 1.14.14.2) and gulonolactone dehydrogenase (EC 1.1.3.8) activities in microsomal preparations and nuclear envelopes were comparable, and an increase in nuclear envelope o-chloroaniline hydroxylase (EC 1.14.14.1) activity twice that occurring in the microsomal fraction was noted (Ichikawa & Mason, 1973). Kasper (1974) records an induction of aryl hydroxylase by 3-methylcholanthrene of 16 times in nuclear envelopes of rat liver and 9 times in the microsomal fraction, indicating that a substantial induction in both membrane types can occur, and the present results show that an even greater induction of UDP-glucuronyltransferase activity in both membranes can be achieved in embryonic-chick liver. The presence of the transferase (EC 2.4.1.17) in chick liver microsomal preparations and its inducibility from scarcely detectable activity to above the mature activity in chick liver in ouo by phenobarbital has been previously established (Wishart & Dutton, 1975). This enzyme is also present in adult rat liver, and its distribution in cellular subfractions parallels that of glucose 6-phosphatase, with 64-76% in the microsomal, 6-974 in the mitochondria1 and 1619% in the nuclear fractions (AmarCostesec et al., 1974). In this type of study it is difficult to be sure that the activity in the non-microsomal fractions is not due to microsomal contamination, and similar comments apply to the finding of low specific activities (compared with the microsomal) of the enzyme in Golgi apparatus and plasma membranes (Nyqvist & M o d , 1971 ; von Bahr et al., 1972). In the present investigation both the nuclear and nuclearenvelope preparations were characterized by electron-microscope morphometry to indicate the microsomal contamination experienced. The absence of suitable marker enzymes for microsomal material in nuclear and nuclear-envelope fractions makes morphometry advisable, and the tendency of the nuclear envelope to form microsomelike vesicles is likely to give a gross overestimate of contamination if only the nuclearenvelope fraction is characterized (see Fry, 1976). To obtain readily detectable quantities of the transferase the livers of phenobarbitaltreated chicks were used. The phenobarbital (1Omg per egg) was injected into the airspace of 17-day embryonated White Leghorn eggs, and the embryo livers were removed on day 20. Nuclei and nuclear envelopes were prepared by the procedure of Kay et al. (1972), except that 2-mercaptoethanol was omitted from the nuclear-digestion media. This procedure purifies the nuclei by a high-density-sucrose spin and subsequent 0.25m~-sucrose wash, and the envelopes are liberated from the washed nuclei by mild deoxyribonuclease digestion at low salt concentration and alkaline pH. It gives a 5 0 4 0 % yield of envelopes with pore complexes recognizable in large sheets of membrane. The microsomal fractions were isolated as described by Burchell et al. (1974) from the first two 700g supernatants, and assayed at the same time as the washed nuclei and nuclear envelopes. Part of the microsomal fraction was subjected to the same digestion procedure as the nuclei. Transferase was assayed as described by KO et al. (1 967).