Effects of cimaterol, a beta-adrenergic agonist, on lipid metabolism in rats.

Rats fed a high carbohydrate diet containing 10 or 100 ppm cimaterol for 4 weeks gained 41% to 59% less fat and 70% to 76% more protein than controls, with no major changes in either energy gain or efficiency of energy retention. Effects of cimaterol on lipid metabolism in these rats were assessed. Cimaterol stimulated lipolysis in vivo and in vitro, but failed to influence rates of de novo fatty acid synthesis in either liver or white adipose tissue. Activities of fatty acid synthetase and malic enzyme in these tissues were also unaffected by cimaterol. Cimaterol administered in vivo failed to affect lipoprotein lipase activity in white adipose tissue, but elevated enzyme activity 67% to 75% in the extensor digitorium longus muscle. Lipoprotein lipase activity in the extensor digitorum longus muscle was also elevated by 66% during a 2 hour incubation with 1 mmol/L cimaterol. We conclude that cimaterol selectively stimulates both lipolysis in white adipose tissue and lipoprotein lipase activity in skeletal muscle, to direct energy away from adipose tissue deposition toward skeletal muscle accretion.

Effects of cimaterol, a beta-adrenergic agonist, on protein metabolism in rats.

Fractional accretion rates of total body 3-methylhistidine containing proteins (actin and myosin) were elevated 40% to 120% in rats fed a high-carbohydrate diet containing 10 or 100 ppm cimaterol for 1 week. Fractional degradation and fractional synthesis rates of these proteins were examined by measuring total body 3-methylhistidine content and urinary excretion of 3-methylhistidine. Consumption of the diet containing 100 ppm cimaterol for 1 week caused a 25% reduction in fractional degradation rates and a concomitant 32% increase in fractional synthesis rates of 3-methylhistidine containing proteins. Effects of cimaterol on fractional accretion, degradation, and synthesis rates of 3-methylhistidine containing proteins diminished after 1 week. Cimaterol failed to influence plasma insulin, triiodothyronine, or corticosterone concentrations. The dramatic increase in accretion of 3-methylhistidine containing proteins observed during the first week rats are fed diets containing cimaterol is caused by reciprocal action on protein degradation and synthesis.

Determination of occupancies of the SPH and GT-IIC transcription factor binding motifs in SV40: evidence for two forms of transcription elongation complex.

Occupancies of the SPH and GT-IIC sequence motifs in the native SV40 late transcription elongation complex were determined by assessing blockage to restriction enzyme cleavage. Cleavages specific to the transcription elongation complex were quantified by radioactive extension labeling and polymerase run-off analysis. The SPH motif was assayed by Sphl digestion and found to be unoccupied. In contrast, digestion with Pvull at the GT-IIC site was blocked in 36% of the complexes, indicating that approximately a third of the complexes are occupied by factor. This fractional occupancy indicates that there are at least two forms of SV40 late transcription elongation complexes, one form with the GT-IIC site occupied by a factor and another with the site vacant.

RNA polymerase locations in the simian virus 40 transcription complex.

Transcription complexes of simian virus 40 can be isolated from cells late in infection in a form that retains the ability to continue transcription in vitro. These complexes have been investigated previously to gain information about the nucleoprotein structure of a transcribing gene. However, several studies have reported that the RNA polymerase molecules in such complexes are located almost entirely at the 5' end of the transcription unit. This would indicate that these complexes cannot be those which were in the process of transcribing mRNA in the cell, because the transcribing complexes would be expected to contain polymerase that were distributed throughout the transcription unit at the time of extraction. Since this issue is important to the interpretation of studies which characterize the extracted complexes, we have determined the polymerase locations using two new approaches. The first employs a new application of a procedure in which the transcription complex DNA is radioactively tagged by transcription in vitro. The second is a new method, which analyzes run-off transcripts generated in vitro. Both methods demonstrate that the polymerases are distributed throughout the genome in the transcription complexes. This result indicates that these complexes were in the process of transcribing mRNA at the time of cell lysis.

Chromatin structure and factor site occupancies in an in vivo-assembled transcription elongation complex.

The chromatin structure specific to the SV40 late transcription elongation complex as well as the occupancy of several sites that bind transcription factors have been examined. These features have been determined by assessing blockage to restriction enzyme digestion. Cleavage specific to the elongation complex has been quantified using ternary complex analysis. This method involves radioactively labeling the complex by in vitro transcription followed by determining the extent of linearization by electrophoresis in an agarose gel. It was found that not only is the origin region devoid of nucleosomes, but there is also no stable factor occupancy at the BglI, SphI, KpnI and MspI restriction enzyme sites within this region. Thus these sites were cleaved to a high degree, meaning that the binding sites for a number of transcription factors, including OBP/TEF-1, TBP, DAP, as well as a proposed positioned nucleosome, are unoccupied in the native viral transcription elongation complex. The absence of these trans-acting factors from their respective binding sites in the elongation complex indicates that they bind only transiently, possibly cycling on and off during the transcription cycle. This finding implies that various forms of transcription complex are assembled and disassembled during transcription and thus supports a 'hit-and-run' model of factor function.