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DNA-dependent RNA polymerases were extracted from rat uterine tissue, partially purified and resolved by DEAE-Sephadex chromatography. RNA polymerases I, II, IIIA, and IIIB eluted at the characteristic ammonium sulfate concentrations of 0.15, 0.28, 0.34, and 0.42 M, respectively. The sensitivity of each peak of polymerase to alpha-amanitin was examined and was shown to be essentially identical to the three classes of RNA polymerases in other mammalian systems. RNA polymerase I was insensitive to high levels of alpha-amanitin, RNA polymerase II was sensitive to low concentrations of alpha-amanitin (50% inhibition at 0.006 mug/ml) and RNA polymerases IIIA and IIIB were sensitive to high concentrations of alpha-amanitin (50% inhibition at 18 mug/ml). The alpha-amanitin sensitivity curve of total RNA synthesis measured in isolated nucleo demonstrated that the activity of each class of RNA polymerase could be quantitated in uterine nuclei. Thus the initial decrease in activity at low concentrations of alpha-amanitin (50% inhibition at 0.005 mug/ml) was attributed to the inhibition of RNA polymerase II activity, the second decrease in activity at higher concentrations of alpha-amanitin (50% inhibition at 15 mug/ml) was attributed to the inhibition of RNA polymerase III activity, and the activity which was resistant to the highest alpha-amanitin concentration tested was attributed to RNA polymerase I activity. When estradiol was given to immature rats 6 h before killing both RNA polymerases I and III levels in nuclei were increased significantly over the control values. The time course of these changes demonstrated that the increases in RNA polymerases I and III were first evident between 1.5 and 3 h following hormone treatment. Significantly, these increases in polymerase I and III in nuclei parallel the published increases for rRNA and tRNA synthesis following hormone treatment. However, the amount of RNA polymerase I and III was not altered upon extraction, suggesting that these changes are due to the alteration in chromatin template activity. Both estradiol and estriol produced identical increases in uterine RNA polymerase I and III 6 h after treatment.
The development of the Multidimensional Health Locus of Control scales is described. Scales have been developed to tap beliefs that the source of reinforcements for health-related behaviors is primarily internal, a matter of chance, or under the control of powerful others. These scales are based on earlier work with a general Health Locus of Control Scale, which, in turn, was developed from Rotter's social learning theory. Equivalent forms of the scales are presented along with initial internal consistency and validity data. Possible means of utilizing these scales are provided.
The peptide hormone, prolactin, when added to organ explants of rat mammary gland, rapidly (within 1 h) induced the accumulation of casein mRNA. Casein mRNA sequences, as determined by hybridization with a specific cDNA probe, were shown to increase for up to 48 h after prolactin addition. The magnitude of this response was dependent upon the day of pregnancy at which the tissue was placed in culture. Maximal levels of induction (as great as 45-fold) were obtained using tissue from 15-day pregnant rats. Further data indicate that two steroid hormones, hydrocortisone and progesterone, were able to modulate the prolactin-induced accumulation of casein mRNA. The continuous presence of hydrocortisone was not necessary for prolactin induction of casein mRNA. However, the presence of hydrocortisone was required for maximal accumulation of casein mRNA. The induction of casein mRNA by prolactin was inhibited in a dose-dependent manner by the simultaneous addition of progesterone to the organ culture. Thus, hydrocortisone appears to potentiate the prolactin induction of casein mRNA, whereas progesterone is able to prevent casein mRNA accumulation. Since mammary gland organ culture is performed in a serum-free, chemically defined medium, this system allows a detailed examination of the mechanims by which a peptide hormone regulates the rapid accumulation of a specific mRNA.
The coat protein of tobacco mosaic virus forms numerous aggregates, including the small A-protein, the disk, and two helical forms. The structures of the disk, the helical protein forms, and the virus are compared. Most of the differences are in the conformation of the chain between residues 89 and 113, which lies in the region of protein at the center of the virus, inside the RNA. It is disordered in the disk, but has a fixed conformation in the virus and the protein helices. The differences between the virus and the two helical protein forms are largely in the conformations of arginines and carboxylic acids in this region.
Transcription of Ad2 DNA templates in the presence of crude cellular extracts supplemented with exogenous (purified) RNA polymerase II is selectively and accurately initiated at the major late viral promoter at map position 16.45. Specific initiation has been demonstrated by a combination of hybridization, nuclease S1 mapping, size and partial sequence (fingerprint) analyses of the transcripts generated with various templates. With intact Ad2 DNA, transcription is terminated ell before the end of the 28 kb transcription unit is reached. With truncated templates (which contain intact promoter regions and several hundred base pair segments of the transcribed region) the expected run-off products are observed, along with a low level of prematurely terminated transcripts. The 560 nucleotide run-off product of the Sma l-f template (coordinates 11.6-18.2) was shown to contain all the large RNAase T1 oligonuc eotides that are characteristic of the corresponding in vivo transcript from this region; in addition, the 5 terminal undecanucleotide appears to be both capped and methylated. We have investigated various parameters (salt, metal ion and template concentrations) that affect the level of specific transcription in the crude system and have found that, under optimal conditions, specific transcription of Ad2 DNA continues for several hours. In addition, specific transcription initiation at the late promoter is observed with extracts derived from either virus-infected or uninfected KB cells and with class II RNA polymerases isolated from either human calf, murine or amphibian cells. RNA polymerase II from wheat germ does not function in this system.
The mechanism by which prolactin, a peptide hormone, regulates casein gene expression has been studied in mammary gland organ culture. After prolactin addition, a 2-4 fold increase in the rate of casein mRNA transcription was observed within 1 hr and maintained for at least 24 hr. This increased rate of transcription is not sufficient to account for the mass accumulation of casein mRNA. The half-life of casein mRNA is also increased 17-25 fold in the presence of prolactin. This change in casein mRNA half-life, coupled with a 2-4 fold increase in the rate of transcription, can account for the normal accumulation of casein mRNA observed after prolactin addition. This hormone-induced change in casein mRNA half-life appeared to be selective, since prolactin was found to exert only a slight effect (1-4 fold) on the half-life of poly(A) RNA determined under identical pulse-chase conditions. The hormonal regulation of casein gene expression thus does not app-ar to be an "all or none" process occurring only at the transcriptional or post-transcriptional levels, but rather may involve a coordinated response at several levels to permit the efficient expression of specialized differentiated functions.
The virus-associated (VA) RNAI gene in human adenovirus 2 DNA has been shown by Wu (Wu, G. J. (1978) Proc. Natl. Acad. Sci. U. S. A. 75, 2175--2179) to be transcribed by RNA polymerase III in a human KB cell-free extract. In the present report we have examined the fidelity of transcription of adenovirus 2 DNA and Xenopus oocyte 5 S DNA templates by RNA polymerase III in extracts derived from cultured human, murine, and amphibian kidney cells, Size and sequences analysis of the discrete transcripts synthesized in these homologous and heterologous systems indicate that they result from accurate transcription of the corresponding genes. The specific transcripts identified include both the adenovirus VA RNAI and VA RNAII, Xenopus 5 S RNA, and VA RNAI and 5 S RNA species with elongated 3' termini. The extracts derived from the various cell types differ in the ability to discriminate between the two VA RNA genes or between the heterogeneous 5 S RNA genes in the cloned DNA fragment. Wherease the human cell extracts transcribe the VA RNAI and VA RNAII genes of adenovirus at a relative frequency close to that observed in isolated nuclei, the amphibian cell extract appears to transcribe only the VA RNAI gene. The amphibian cell extract transcribes primarily that 5 S RNA gene (within 5 S DNA) which encodes the dominant oocyte 5 S RNA, whereas the human cell extract transcribes at least two distinct 5 S RNA genes. Additionally, it is shown that the VA RNAI and VA RNAII genes have separate promotor sites. The kinetics of the transcription reactions have been examined and conditions optimal for specific transcription have been established by examining the effects of salt, metal ion, and template concentrations on both total and specific RNA synthesis. It is also shown that components in the cell-free extract (from human cells) are active in directing the accurate transcription of adenovirus DNA by purified RNA polymerase III.
Locus of control, an individual difference construct from social learning theory, has shown some promise in predicting and explaining specific health-related behaviors. Research is reviewed on the utility of the locus of control construct in understanding smoking reduction, birth control utilization, weight loss, information-seeking, adherence to medication regimens, and other health or sick-role behaviors. Implications for health educators are presented.