Measurement of the rate and velocity of movement by single heart cells in culture.
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Biomedical subjects
Publications and source records attributed to S H Wilson.
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A DNA polymerase activity that promotes the synthesis of poly(dT) has been found in association with intracisternal A-type particles isolated from several mouse tumors. The poly(dT) synthesis activity requires a DNA or RNA primer, is optimal at high salt concentration, prefers magnesium over manganese, and is stimulated by poly(rA). No significant incorporation of dAMP, dGMP, or dCMP was detected in the presence of several RNA and DNA template-primers. The enzyme activity differs in several of its properties from the poly(rA)-directed DNA polymerase activity associated with Rauscher murine leukemia virus.
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The rates of synthesis of ribosomes, 5S RNA, and tRNA necessary to maintain the steady-state concentrations of these entities in liver cytoplasm of adult rats were determined. On the average, each liver cell in the adult rat synthesizes 650 ribosomes, 650 molecules of 5S RNA, and 11,000 molecules of tRNA each minute. The numbers of genes per liver cell for rRNA, 5S RNA, and tRNA were 330, 1660, and 13,000, respectively, as determined by RNA: DNA hybridization experiments. Thus, on the average, individual genes for rRNA, tRNA, and 5S RNA are transcribed twice a minute, once a minute, and once every 2.5 minutes, respectively, in the adult rat liver.
Starvation of rats for several days led to marked decrease in cytoplasmic polysomes and accumulation of breakdown products having S values less than 200s. Re-feeding of the starved animals induced a rapid reassembly of polysomes. These newly formed polysomes, in the presence of actinomycin D, decayed in a biphasic fashion: about two-thirds decayed with an apparent half-life of 3-3(1/2)hr. but the other one-third were much more stable. Evidence that polysome decay is an accurate reflexion of messenger RNA stability is presented, and it is concluded that in the presence of large doses of actinomycin D, rat-liver cytoplasm contains messenger RNA classes of widely varying stability, the more stable class having a half-life of at least 80hr. The half-life of liver ribosomes was also determined and was found to be 110-127hr.
Certain qualitative aspects of protein synthesis in the livers of starved, starved-re-fed and actinomycin D-treated rats have been examined by polyacrylamide-gel electrophoresis. Animals were exposed to a mixture of (14)C-labelled acids for 18-20min. and killed, and an ultrasonic extract of newly formed protein in microsomal vesicles was prepared and examined by gel electrophoresis. In normal and starved-re-fed animals, 27% of the newly synthesized protein was albumin. During starvation, when RNA synthesis was decreased, the percentage of newly formed protein as albumin rose. After actinomycin D treatment of starved-re-fed rats, when only stable messenger RNA persisted in the cytoplasm, albumin synthesis increased to 63% of the total. This finding suggested that albumin was the primary protein synthesized on stable messenger RNA.
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The mammalian DNA beta-polymerase (beta-pol) gene is constitutively expressed in cultured cells as a function of growth stage and DNA replication, but is expressed in rodents in a tissue-specific fashion. As revealed by transient expression experiments with wild-type and mutated beta-pol promoter fusion genes, the cloned human beta-pol promoter is transcriptionally regulated by signals acting through the single palindromic sequence (GT-GACGTCAC) known as an ATF/CRE-binding site centered at position -45 in the core promoter. Although the mere presence of the ATF/CRE palindromic sequence in a promoter does not always confer cAMP responsiveness or protein binding over and around the ATF/CRE sequence, we find that agents that increase cAMP levels (forskolin and IBMX) in HeLa cells activate the beta-pol promoter; activation also can be observed by coexpression of the protein kinase A catalytic subunit. Experiments with mutagenized beta-pol promoters indicate that the ATF/CRE-binding site mediates these effects. Thus, the ATF/CRE-binding site in the context of this TATA-less constitutive promoter is able to respond to the kinase A signal transduction pathway.