Albumin extinction followed by de novo methylation of its gene in somatic hybrids of a rat hepatoma

We have previously identified an Msp I site at the 5′ end of the rat albumin gene whose undermethylation is necessary but not sufficient for stable albumin expression in rat hepatoma cells [1]. We have also shown that the extinction of albumin expression in somatic hybrids is not the result of methylation at this site, since for two different crosses, rapid extinction was found to occur in the absence of any de novo methylation of the previously active gene[2]. In the present study, we examine albumin expression and albumin gene methylation for independent hybrid clones isolated from crosses between albumin expressing rat hepatoma cells and cells of two different non-expressing lines. The cells from hybrid clones of both crosses are characterized by stable extinction of albumin expression. Moreover, we find that de novo methylation of the “extinguished” albumin gene can occur in somatic hybrids, but only some weeks after the gene has ceased to be expressed.     

1H-NMR study of the lambda operator site OL1: assignment of the imino and adenine H2 resonances.

One- and two-dimensional proton NMR methods are being used to study the synthetic lambda operator site O-L1, a 17 base-pair DNA duplex recognized by lambda repressor and Cro protein. The complete assignment of the 17 imino protons, which participate in Watson-Crick hydrogen bonding, and of the eight adenine H2 protons, which lie in the minor groove of the double helix, is presented.     

Nucleotide sequence of the VP1 gene of the foot-and-mouth disease virus strain A Venceslau

The VP1 coat protein of FMDV strain A Venceslau (Aven) consists of 213 amino acid residues. Serum neutralization tests demonstrated that strain Aven is closely related to strain A Argentina/79 (A₇₉) but significantly different from strain A₂₄ Cruzeiro (A₂₄). There is a strong correlation between the amino acid sequences and the serological data. Nucleotide and amino acid sequence analyses of VP1 showed that serologically related viruses (Aven and A₇₉) differ less in this region of the genome than those of serologically distinct viruses (Aven vs. A₂₄). The most significant variation between Aven and A₂₄ occurs at amino acid positions 43 to 46, in which all four residues are different.     

Effect of treatment with the MER tubercle bacillus fraction on syngeneic plasma cell tumors of BALB/c mice

Summary MER administered prophylactically prolonged the survival of BALB/c mice challenged with transplants of syngeneic plasmacytomas to a moderate but significant extent. In contrast, MER exerted little therapeutic action when given alone at or after tumor implantation.Combined treatment, with MER introduced prophylactically and cyclophopshamide (CY) after tumor implantation, decreased the incidence of recurrence of one of the tumors tested (MPC-11 NP, a non-myeloma protein producer) and prolonged host survival significantly as compared with animals subjected to CY therapy alone. When, instead, MER was introduced at the time of tumor challenge or thereafter to animals also treated with CY, the therapeutic response was not appreciably different from that of mice under therapy with CY only. With regard to the second plasmacytoma (MPC-11 P, a myeloma protein producer), mice treated with CY and given MER prior to or after challenge showed similar responses to animals given chemotherapy only; when MER was injected at the time of challenge and CY thereafter, the chemoimmunotherapy was somewhat inferior to chemotherapy alone.This work was supported by US Public Health Service Contract NO1-CM-12127, and by grants from Mrs. J. H. Hazen, Ruth Estrin Goldberg Memorial for Cancer Research, Leukemia Research Foundation, Inc., Concern Foundation, Inc., and Mr. and Mrs. M. Gordon     

Serum relaxin levels in prostaglandin E2 induced abortions

Relaxin, a peptide hormone produced only by the corpus luteum of pregnancy, can be used as a marker of luteal function in human pregnancy. Serum immunoreactive relaxin levels were measured serially in six women having second trimester abortions induced with intravaginal prostaglandin E2 (PGE₂) suppositories. All patients aborted within 17 hours of the first suppository. No significant changes were detectable in serum relaxin levels in any of the patients. It is concluded that PGE₂ does not interfere with the corpus luteum's ability to secrete relaxin in the second trimester of human pregnancy.     

Control of arginine utilization in Neurospora.

The response of Neurospora to changes in the availibility of exogenous arginine was investigated. Upon addition of arginine to the growth medium, catabolism is initiated within minutes. This occurs prior to expansion of the arginine pool or augmentation of catabolic enzyme levels. (Basal levels are approximately 25% of those found during growth in arginine-supplemented medium.) Catabolism of arginine is independent of protein synthesis, indicating that the catabolic enzymes are active but that arginine is not available for catabolism unless present in the medium. Upon exhaustion of the supply of exogenous arginine, catabolism ceases abruptly, despite an expanded arginine pool and induced levels of the catabolic enzymes. The arginine pool supports protein synthesis until the cells regain their normal capacity for endogenous arginine synthesis. These observations, combined with the known small level of induction of arginine catabolic enzymes, non-repressibility of most biosynthetic enzymes, and vesicular localization of the bulk of the arginine pool, suggest that compartmentation plays a significant role in controlling arginine metabolism in Neurospora.     

Chromium in biological systems, I. Some observations on glucose tolerance factor in yeast

Glucose tolerance factor (GTF) has been isolated from a commercially available yeast extract powder, by a simple procedure under mild conditions. This cationic yellow material enhances considerably CO2 production in several yeast strains, after a lag time which can be eliminated by preincubation with glucose. The enhancement of CO2 production by GTF is not specific for glucose, and its effect on galactose raises the possibility that it influences the transport of the sugar to the cells. The ineffectiveness of GTF on cell free extract and the results of a Michaelis plot for CO2 production support this hypothesis.     

ATP synthesis catalyzed by the ATPase complex from Rhodospirillum rubrum reconstituted into phospholipid vesicles together with bacteriorhodopsin

The coupling factor ATPase complex extracted by Triton X-100 from the photosynthetic bacterium Rhodospirillum rubrum could be incorporated into phospholipid vesicles after removal of the Triton. Vesicles reconstituted with this F₀ · F₁-type ATPase together with bacteriorhodopsin were found to catalyze, in the light, net ATP synthesis which was inhibited by the energy transfer inhibitors oligomycin and N,N-dicyclohexylcarbodiimide as well as by uncouplers. In vesicles reconstituted with the crude ATPase up to 50% of the observed rate of phosphorylation was independent on light and bacteriorhodopsin and insensitive to the above-listed inhibitors. This dark activity was, however, completely blocked by the adenylate kinase inhibitor, p¹,p⁵-di(adenosine-5′)pentaphosphate, which did not affect at all the net light-dependent phosphorylation nor the ATP-³²P_i exchange reaction. Vesicles reconstituted with the purified ATPase catalyzed only the light- and bacteriorhodopsin-dependent diadenosine pentaphosphate-insensitive phosphorylation. The rate of this photophosphorylation was found to be proportional to the amount of ATPase and bacteriorhodopsin, and linear for at least 20 min of illumination. These results indicate that the purified ATPase contains the complete assembly of subunits required to transduce electrochemical gradient energy into chemical energy.     

Prevention by the MER tubercle bacillus fraction of immunosuppression induced by cancer chemotherapeutic agents

Summary Contact hypersensitivity (CH) to 2,4-dinitro-1-fluorobenzene (DNFB) was induced in guinea pigs and mice by DNFB skin application. Development of CH was suppressed in both species either by cyclophosphamide (CY) treatment after sensitization or by single intravenous injection of dinitrobenzene-sulfonate (DNBS) before sensitization (hapten-induced tolerance). Additional treatment schedules were employed in guinea pigs, with the following results: Suppression of CH by injection of DNBS concomitant with sensitization; abrogation of hapten-induced tolerance by administration of CY before sensitization; and potentiation of CH skin reactivity by administration of CY before sensitization.Pretreatment by two injections of the methanol extraction residue (MER) tubercle bacillus fraction restored significantly the ability of CY treated animals to respond to DNFB sensitization. In contrast, administration of MER either by one injection before sensitization, concomitant with DNFB, or after sensitization did not prevent immunosuppression by CY.MER treatment was not effective in reversing hapten-induced tolerance in mice, and had only an occasional effect on this process in guinea pigs. Abrogation of hapten-induced tolerance and potentiation of DNFB sensitization by CY in guinea pigs were also not influenced by MER treatment.Supported by Contract NO1-CM-12127 from the NCI and by research grants from Concern Foundation, Inc., the Lautenberg Endowment, the National Council for Research and Development, Israel, and the GSF Munich, Germany, and the Leukemia Research Foundation, Inc.