Phenobarbital stimulation of housefly chromatin template activity

The effect of phenobarbital on the chemical composition and template activity for Escherichia coli RNA polymerase of housefly chromatin is examined. Chromatin isolated from the resistant Fc housefly strain after administration of phenobarbital has a much higher template activity as compared to control chromatin. The effect is minimal in a susceptible housefly strain. Phenobarbital treatment alters the composition of the RNA polymerase product which results in an increase of the $\text{(A + U)}\text{(C + G)}$ ratio with chromatin from the Fc strain as template, while the $\text{(A + G}\text{)}\text{(}\text{C + U)}$ and $\text{(A + U}\text{)}\text{(}\text{C + G)}$ ratios are decreased when chromatin of the susceptible strain is used.A substantial portion of the intracellular phenobarbital is physically bound to nuclei and microsomes, but most of it is associated with cytosolic macromolecules. Less than 0.2% is covalently bound to cellular macromolecules. Phenobarbital apparently binds to a small mol. wt cytosolic macromolecule(s) as shown by sucrose density gradient centrifugation and Sephadex G-25 column chromatography.     

Repressed template activity of chromatin of pea roots treated by aluminium

To determine the mechanism of aluminium toxicity in the pearoot, we investigated both the in vivo and in vitro effectsof aluminium on chromatin. The ratio of histone or non-histoneproteins to DNA in chromatin was slightly increased by aluminiumtreatment. However, chromatin with an rf of 0.086 (rf: molarratio of bound aluminium to DNA-phosphate) prepared from aluminiumtreated roots had template activity for RNA synthesis that washalf of the control. The template activity of the chromatintreated with aluminium in vitro decreases with increasing rfvalues. In vitro treatment of chromatin with aluminium altered the spectrophotometricproperties of chromatin, including the shift up in the absorptionof the region beyond 320 nm and the decrease in the ratio ofmaximum to minimum absorption, with increasing concentrationsof treated aluminium. The dialysis of DNA or histone againstan aluminium solution indicates that scarcely any binding ofaluminium to histone occurred, but the binding of aluminiumto DNA took place rapidly. ¹Nippon Shinyaku Co. Ltd., Minami-ku, Kyoto 601, Japan. (Received June 20, 1980; )     

RNA chain elongation on a chromatin template.

The rate of RNA chain elongation has been measured with DNA and chromatin as template. RNA propagation on chromatin is about 50% of the rate found with DNA. Kinetic experiments demonstrate that the inhibition is not due to interference with the addition of the nucleoside triphosphates. Analysis of the dependence of propagation on the Tm of DNA shows that the chromatin proteins interfere with the translocation of the RNA polymerase along the DNA template.     

Increased template activity in chromatin from cadmium chloride treated pea tissues

Increases in the synthesis of the isoflavonoid, pisatin, and the activity of phenylalanine ammonia lyase (PAL) are induced in excised pea pods by low concentrations (5×10^?4M) of CdCL₂. The induction of pisatin synthesis and PAL are suppressed if RNA-synthesis-inhibiting concentrations of 6-methyl purine, actinomycin D or ∝-amanitin are applied within 1 h of inducer application. Cycloheximide (0.1 mg/ml) blocks the induction of these responses if applied to tissues within 6 h after inducer application. Within 1 h after CdCl₂ (5×10^?4M) is applied to pods there is an increase in the rate of synthesis of all sizes of RNA as well as an increase in the template activity and dye binding capacity of pea chromatin. The results support the hypothesis that conformational changes in DNA are associated with the induction process.     

Modification of rat liver chromatin by N-methyl-N'-nitro-N-nitrosoguanidine or N-ethyl-N'-nitro-N-nitrosoguanidine and template activity for RNA synthesis by Escherichia coli RNA polymerase after reconstitution.

Rat liver chromatin was fractionated into DNA, histones and non-histone chromosomal proteins and each component was modified with N-methyl-l-N'-nitro-N-nitrosoguanidine of N-ethyl-N'-nitrosoguanidine. The radioactivity of 14C-labeled alkyl or guanidino moieties of both compounds bound significantly to both histones and non-histone chromosomal proteins and the binding of N-methyl-N'-nitro-N-nitrosoguanidine was higher than N-ethyl-N'-nitro-N-nitrosoguanidine. However the binding of both compounds to DNA was very low and its significance was hard to evaluate. All of the three components, one of which was modified, were reconstituted into chromatin, then, [3H]UMP incorporation into acid insoluble material using Escherichia coli RNA polymerase (EC 2.7.7.6) was measured. Only with the reconstituted chromatin containing histones modified either by N-methyl-N'-nitro-N-nitrosoguanidine or N-ethyl-N'-nitro-N-nitrosoguanidine, the template activity increased drastically; i.e., about 10 or 5 times higher than that with the unmodified reconstituted chromatin, respectively. However, any remarkable alteration in the electrophoretic pattern of protein fraction of the reconstituted chromatin could not be found. The results obtained in this study are discussed in the context that the modified histones could give rise to change in the mutual interaction of chromosomal components during the reconstitution of chromatin accompanied with the increase of chromatine template activity.