Prolactin (PRL) gene expression in three strains of GH cells (rat pituitary tumor cells) has been quantitated by measurement of : (a) intracellular and extracellular PRL, (b) cytoplasmic translatable PRL-specific mRNA (mRNAP R L), and (c) molecular hybridization of cytoplasmic poly(A) RNA to cDNA PRL (DNA complementary to mRNA PRL) . Three GH cell lines utilized in this investigation were a PRL-producing (PRL + ) strain, GH 4C,, a PRL nonproducing 5-bromodeoxyuridine resistant (PRL -BrdUrd') strain, F,BGH,2C,, and a new strain, 928-9b, derived by fusion of PRL + cells with a nuclear monolayer of the PRL-, BrdUrd r GH cell strain . PRL production is a characteristic of 928-9b cells, but the level of PRL production (2-4 Ftg/mg protein/24 h) is much lower than that of the PRL + strain, GH 4C, (15-25 ,ug/mg protein/24 h) . Levels of cytoplasmic translatable mRNA PRL and cytoplasmic PRL-RNA sequences quantitated with a cDNAPRL probe were also much lower in 928-9b as compared to the PRL + parent . PRL-RNA sequences could not be detected in the PRL -strain . Thyrotropin-releasing hormone (TRH) stimulates PRL synthesis about threefold and inhibits growth hormone (GH) synthesis 72% in the PRL+ strain . TRH has no effect on the synthesis of either PRL or GH in the 928-9b strain, although TRH receptors could be detected in these cells. Stimulation of PRL synthesis in the PRL+ strain by TRH could be correlated with increases in levels of cytoplasmic translatable mRNAPRL and increases in cytoplasmic PRL-RNA sequences. These results demonstrate that the graded expression of the PRL gene at the basal level, and in response to TRH, is caused by the regulated production of specific mRNA, i.e ., mRNA PRL in these three GH cell strains.Different clonal strains of rat pituitary tumor cells (GH cells) in culture produce different amounts oftwo protein hormones, prolactin (PRL) and growth hormone (GH). Certain properties of the GH strains used in this investigation are described in Table I. The GH strains differ from each other not only in their basal level production of PRL but also in their response to the physiological modulator, thyrotropin-releasing hormone (TRH) and to the drug, 5-bromodeoxyuridine (BrdUrd) (l, 2). The GH,C, and GH3 subclones respond to TRH with a stimulation ofPRL synthesis and an inhibition of GH synthesis (for review, see reference 13.). However, GH12C1 and the BrdUrd' (BrdUrd resistant) derivative of GH12C1 , the F,BGH12C, substrain, do not produce any detectable PRL and do not respond to TRH . We have previously reported that PRL synthesis can be induced in these two PRL-nonproducing (PRL-) strains by treatment with the drug BrdUrd (1) . These results suggest that, in these cells, PRL synthesis is under the influence of a rigid cellular control mechanism/s which does 6 not permit PRL gene expression, but which is affected by incorporation of the drug into the DNA, subsequently permitting the PRL gene to be expressed .The varied basal level expression of the PRL gene in these GH strains and the ...
Laboratory in vitro testing of various remedies from the Old English Leechbooks and Lacnunga does not support previous assertions that Anglo-Saxon medical remedies would have been efficacious. For example, the remedy for a stye in the eye takes ingredients that individually have anti-bacterial properties and compounds them into a mixture with no effect on common bacteria. We conclude that Anglo-Saxon remedies were not likely to have cured the ailments for which they were prescribed and that researchers, rather than asserting the probable prowess of the Anglo-Saxon læce, should instead focus on what people in the time period believed would have helped them.
Prolactin-specific RNA (RNApRL) in total nuclear RNA and in cytoplasmic poly(A)+RNA isolated from GH (rat pituitary) cells was selectively hybridized to immobilized cloned cDNApRL. Agarose gel electrophoresis. of the nuclear RNAPRL sequences eluted from the nitrocellulose filters revealed several RNA species of approximately 25-30, 18-19, and 12-13 S. Only the 12-13 S RNA species could be detected in the cytoplasmic poly(A)+RNA fraction. Comparative analysis oftotal nuclear RNA of control and thyrotropin-releasing hormone (thyroliberin)-treated cells by the reverse Southern blot technique demonstrated increased levels of all the nuclear RNAPRL species in hormone, treated cells. Nuclear and cytoplasmic RNAPRL sequences in control and treated cells were quantitated by molecular hybridization to clonedcDNApRL. The 2-to 34old stimulation ofPRL production by thyrotropin-releasing hormone-treated GH4C1 cells could be correlated to the. corresponding increase of nuclear RNAPRL sequences. The hybrid strain, which produces 1/5th the amount of PRL that the parent GH4Cj does, had 1/5th the amounts of nuclear RNAPRL sequences. Thyrotropin-releasing hormone affected neither prolactin production nor nuclear RNAPRL level in 928-9b cells. RNAPRL sequences could not be detected either in nuclei or in cytoplasm of prolactin nonproducing F1BGH12C, cells. However, prolactin production could be induced and RNAPRL sequences could be detected in the total nuclear.RNA and in cytoplasmic poly(A)+RNA fraction after treatment-of this GH cell substrain with 5-bromodeoxyuridine. These results demonstrate that differential basal prolactin production and 'its modulation by thyrotropin-releasing hormone and by 5-bromodeoxyuridine can be correlated to the altered levels'of nuclear RNAPRL sequences in the three GH cell strains.Prolactin (PRL) production in a hybrid strain (928-9b) (8). However, it has not~been known whether the increased PRL production results from increased production of nuclear and cytoplasmic RNAPRL sequences. This investigation also demonstrates the close correlation ofBrdUrd-induced PRL synthesis with a concomitant increase in nuclear and cytoplasmic RNAPRL sequences, from undetectable levels to detectable amounts. MATERIALS AND METHODSGH cells are clonal strains of rat pituitary tumor cells in culture (9). Isolation, growth conditions, and properties of the three substrains used in this investigation have been described in detail (1).Isolation of Nuclear and Cytoplasmic RNA. Isolation of nuclei and cytoplasm and of total RNA and poly(A)+RNA from these subcellular fractions has been described (10,11 by insertion of cDNAPRL (approximately 900 base pairs) into plasmid pBR322 at the Pst I site.
GH12C1, a clonal strain of rat pituitary tumor cells in culture (GH cells), does not produce detectable amounts of prolactin. 5-Bromodeoxyuridine (BrdUrd), the thymidine analogue, at sublethal concentrations (3-5 /xg/ml) induces prolactin synthesis in these cells. BrdUrd also induces prolactin synthesis in F1BGH12C1 cells, a BrdUrd resistant (BrdUrd r) substrain isolated from GH12C1 cells. The F1BGH~2C1 strain is not drug dependent, but its resistance to BrdUrd is a stable phenotype. The significant features of the induction of prolactin synthesis in the BrdUrd r strain are the increased net synthesis of prolactin and the shortening of the lag period of prolactin induction. As BrdUrd concentration in the growth medium is increased, the rise in prolactin synthesis parallels the increased incorporation of BrdUrd into DNA. Prolactin synthesis is first detected when BrdUrd replaces 20-25% of the thymidine in DNA. BrdUrd can replace up to 75-80% of the thymidine within 2 d of treatment. Partial starvation of these cells under specified growth conditions does not affect the general growth pattern of the cells, general protein synthesis, and thymidine uptake, but does affect DNA synthesis. When cells are cultured under conditions in which DNA synthesis is preferentially inhibited, BrdUrd does not induce prolactin synthesis, suggestive of a DNA-mediated mechanism of action for the drug.KEY WORDS 5-bromodeoxyuridine prolactin gene expression 9 rat pituitary tumor ceilsThe drug 5-bromodeoxyuridine (BrdUrd), a thymidine analogue, seems to have diverse effects on the various processes of differentiation in eukaryotic cells. These may be classified into two distinct types. BrdUrd suppresses the synthesis of specific proteins in certain cells, and in other cell types the drug induces the production of certain cell-specific proteins. Rutter et al. (12) described in detail the role of BrdUrd on different eukaryotic cell systems. On the basis of the different effects of this drug, these authors discussed the possible DNA-linked and non-DNA-linked mechanisms of action of BrdUrd.Holtzer and Abbott (7) postulated that BrdUrd selectively inhibits the synthesis of luxury molecules without grossly depressing the synthesis of essential molecules of the cells. Priesler et al. (11) claimed that inhibition of differentiated function J. CELL BIOLOGY 9 The Rockefeller University Press 9
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.