Myocyte and nonmyocyte RNA polymerase activity and chromatin template function.
Myocardial and nonmyocardial cell nuclei were isolated from ventricles of adult male Wistar rats by means of sucrose density centrifugation. RNA polymerase activity in myocyte nuclei was approximately 80-90% higher than in nonmyocytes. Total myocyte chromatin template activity using Escherichia coli RNA polymerase was linear and about onefold greater than the nonmyocyte fraction over a fivefold range of chromatin concentrations. Preincubation time required for RNA polymerase to form a stable binding complex with chromatin was at least 40 min for myocyte and 30 min for nonmyocyte nuclear subsets. Titration of chromatin against a fixed amount of RNA polymerase (5 micrograms) showed that 5 micrograms of myocyte chromatin (as DNA) and 8 micrograms of nonmyocyte chromatin (as DNA), respectively, were required to saturate the enzyme. These results indicate that myocyte chromatin can support greater enzyme binding than can nonmyocyte chromatin. The distribution of DNA fragments from isolated myocyte and nonmyocyte nuclei were similar when alkaline sucrose density centrifugation was used. Chromatin prepared from these nuclear subsets showed no difference in degree of DNA fragmentation. These observations indicate that compared with nonmuscle cells, myocytes from adult rat hearts have higher RNA polymerase activity, greater overall chromatin template function, and higher binding capacity for RNA polymerase. Collectively, these data suggest that adult cardiac muscle cells should have a larger potential for RNA synthesis than nonmuscle cells.
D000818:Animals; D002467:Cell Nucleus / Q000201:enzymology / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002843:Chromatin / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D066298:In Vitro Techniques; D008297:Male; D009206:Myocardium / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D013698:Templates, Genetic
D002843:Chromatin; D004247:DNA; D012321:DNA-Directed RNA Polymerases