Expression of the SOS response following simultaneous treatment with methyl-nitrosoguanidine and mitomycin C in Escherichia coli

Simultaneous treatment of Escherichia coli cultures with methyl-nitrosoguanidine and mitomycin C induces recA-dependent inhibition of respiration but not inhibition of cell division. This pattern of SOS functions expression is the same as that is found following treatment with methyl-nitrosoguanidine alone and contrary to the pattern induced after mitomycin C addition. The same result is obtained when a culture of E. coli RecA441 (formerly tif) is shifted to 42 degrees C and treated simultaneously with methyl-nitrosoguanidine. The suppressor effect of this compound over the pattern of SOS functions expression induced by mitomycin C or high temperature in recA441 mutants is directly related to the increase in its dose. Moreover, the division temperature-sensitive mutant ftsA treated with methyl-nitrosoguanidine and high temperature does not show any decrease in its normal filamentous growth when cultured at 42 degrees C. This indicates that the effect of methyl-nitrosoguanidine on the recA-independent inhibition of cell division is not due to any indiscriminate effect of this compound over the division process. These results suggest that the specific kind of lesion caused in DNA is very important in determining which SOS function is induced.     

Antagonism of diazepam-induced feeding in rats by antisera to opioid peptides

Diazepam-induced feeding in rats is antagonized not only by the opiate antagonist naloxone but also intraventricular administration of specific antisera to the endogenous opioid peptides met-enkephalin or beta-endorphin. Pituitary beta-endorphin is probably not implicated in the diazepam effect since blockade with the glucocorticoid dexamethasone of the release of beta-endorphin from the anterior pituitary does not modify the diazepam-induced feeding, which is however prevented by TRH, a suggested physiological antagonist of some of the effects of opioid peptides. The possible central participation of both beta-endorphin and met-enkephalin in the ingestive behavior induced by diazepam gives further support to the postulated physiological role of endogenous opioids in appetite regulation.     

Inhibition of gastric acid secretion by intracerebral injection of calcitonin gene related peptide in rats

Calcitonin gene related peptide (CGRP), a 37 amino acid peptide recently characterized from the rat brain, injected into the cisterna magna or the lateral hypothalamus, inhibited gastric acid secretion and increased serum gastrin levels in conscious pylorus-ligated rats. CGRP suppressed pentagastrin- and histamine-induced stimulation of gastric secretion in urethane-anesthetized gastric fistula rats. Peptide action was dose-dependent, not modified by adrenalectomy and peripheral sympathectomy induced by guanethidine pretreatment and reversed by vagotomy. These results demonstrate that intracisternal or intralateral hypothalamic injection of CGRP inhibits stimulated gastric acid secretion. The mechanism of action is vagal dependent and unrelated to modification of gastrin secretion or activation of sympathetic outflow.     

Characterization of specific receptors for atrial natriuretic factor in bovine adrenal zona glomerulosa

We have recently shown that synthetic rat atrial natriuretic factor (ANF) directly inhibits mineralocorticoid and glucocorticoid secretion in cultured bovine adrenal cells with a potency of 100 pM. [125I]iodo-ANF was used in the present study to characterize potential receptor sites in bovine zona glomerulosa membranes. ANF binds to a class of high affinity binding sites with a pK of 10.2 and a density of 1.3 pmol/mg protein. Detailed competition curves with ANF document a class of high affinity sites with a pK of 10.2 and also a second class of lower affinity sites with a pK of 8.5. Nonspecific binding amounts to less than 10% of [125I]iodo-ANF binding at concentrations less than 100 pM. High affinity binding of [125I]iodo-ANF is reversible with a half-time of association of 15 minutes at 25 pM and a half-time of dissociation of 140 minutes. Monovalent cations Na, Li and K equipotently enhance [125I]iodo-ANF specific binding. Divalent cations Mg, Ca and Mn also increase [125I]iodo-ANF specific binding, with Mn being the most active cation. No effect of guanine nucleotide could be detected on ANF binding. The binding of [125I]iodo-ANF is very specific and is not inhibited by 1 microM angiotensin II, ACTH, VIP, somatostatin, Leu-enkephalin, dynorphin or by the N-terminal of POMC. The N-terminal fragment ANF-(1-16) is also completely inactive. Reduction of the disulfide bridge of ANF inactivates the peptide. This enabled the development of a highly specific radio-receptor assay for ANF with a minimum detectable dose of 2 femtomoles. The results document the specific receptor involved in the potent inhibitory effect of ANF on adrenal steroidogenesis and indicate that bovine adrenal zonal glomerulosa provide a highly sensitive system for studying the recently discovered atrial natriuretic factor.     

Bidentate peptides: highly potent new inhibitors of enkephalin degrading enzymes

Three series of bidentates bearing an hydroxamic or an N-Acyl-N-hydroxy amino group on structures related to Phe-Gly or Phe-Ala exhibit strong inhibitory potency against purified enkephalinase with IC50 values in the 4 to 15 nM range. As with thiol-containing inhibitors, such as thiorphan, the most active compounds are those in which a methylene spacer separates the benzyl P1' moiety from the Zn coordinating residue. Formation of a bidentate complex with the metal enzyme is clearly demonstrated by a loss of potency of three order of magnitude following the removal of one component of the bidentate group. All the compounds studied are unable to interact with angiotensin converting enzyme (IC50 greater than 10,000 nM). Moreover, compounds of the general formula HONHCO-CH2-CH(CH2 phi)-CONH-CH(R)-COOH belonging to the most active series of enkephalinase blockers (IC50 approximately 4 nM) behave also as highly potent and competitive inhibitors (IC50 approximately 10 nM) of a Tyr-Gly releasing dipeptidylaminopeptidase purified from rat brain. The pure steroisomer [(R)-3-(N-hydroxy)carboxamido-2-benzylpropanoyl]-L-alanine designated kelatorphan, exhibits also a relatively good inhibitory potency against aminopeptidases (IC50 approximately 10 microM) and can be considered as the first virtually complete inhibitor of enkephalin metabolism. This very interesting property of inhibiting all three enzymes of enkephalin metabolism could enhance the required selectivity for a possible clinical use of these inhibitors as new analgesic and psychoactive drugs.     

Effect of mismatched base pairs on the fate of donor DNA in transformation of Streptococcus pneumoniae

Summary Investigation of the mechanism that discriminates against mismatched base pairs in transformation of Streptococcus pneumoniae of genotype hex ⁺ was based on the use of a radioactively labeled cloned fragment of pneumococcal DNA as donor in transformation. The fate of the donor label was followed by lysis of the transformed cells and separation by agarose gel electrophoresis of DNA fragments generated by restriction endonucleases. As a result of Hex action, most of the donor DNA fragment, which was a few kilobases in length, was lost when a mismatched base pair occurred between donor and recipient DNA. This was not observed in hex ^- recipient cells. Kinetic studies of mismatch-induced donor DNA loss showed that the process is faster in strain 800, an R6 derivative, than in DP 1601, a strain of different origin. In the latter strain, the amount of donor label that becomes double stranded rises substantially, indicating extensive formation of donorrecipient heteroduplex structures, before falling to the expected level. At 30°C the process is essentially completed 15 min after entry.     

A temperature-sensitive mutant of Escherichia coli that shows enhanced misreading of UAG/A and increased efficiency for some tRNA nonsense suppressors

Summary A spontaneous mutant was isolated that harbors a weak suppressing activity towards a UAG mutation, together with an inability to grow at 43° C in rich medium. The mutation is shown to be associated with an increased misreading of UAG at certain codon contexts and UAA. UGA, missense or frameshift mutations do not appear to be misread to a similar extent. The mutation gives an increased efficiency to several amber tRNA suppressors with-out increasing their ambiguity towards UAA. The ochre suppressors SuB and Su5 are stimulated in their reading of both UAG and UAA with preference for UAG. An opal suppressor is not affected. The effect of the mutation on the efficiency of amber and ochre suppressors is dependent on the codon context of the nonsense codon.The mutated gene (uar) has been mapped and found to be recessive both with respect to suppressor-enhancing ability as well as for temperature sensitivity. The phenotype is partly suppressed by the ochre suppressor SuC. It is suggested that uar codes for a protein, which is involved in translational termination at UAG and UAA stop codons.     

Germacranolides from Schkuhria anthemoidea

The isolation of eucannabinolide and three new sesquiterpene lactones from Schkuhria anthemoidea is reported. The structures and stereochemistries of the new compounds were established by chemical and spectroscopic means. The structure of santhemoidin B was confirmed by X-ray crystallography.     

An electrogenic proton pump associated with the Golgi apparatus of mouse liver driven by NADH and ATP

Golgi-apparatus membranes, isolated from mouse liver, pump protons inwards, when supplied with NADH or ATP. The acidification of Golgi-apparatus cisternae and vesicles was detected with neutral red, a permeant dye, as a difference in absorbance at 550 nm minus that at 600 nm. The maximum rates detected with NADH and ATP were between 0.0006-0.0009 and 0.0030-0.0050 delta OD units/mg of protein/min, respectively, at pH 7.5. The outside buffer used was a bovine serum albumin suspension. The acidification of Golgi apparatus was inhibited from 45 to 100% by ionophores and from 22 to 100% by uncouplers. The results implicate both ATP and a redox system coupled to NADH oxidation in the acidification of Golgi-apparatus membranes.     

Use of microcalorimetry for the characterization of marine metabolic activity at the water-sediment interface

Metabolic processes occurring at the sea-water-sediment interface were studied using a circulation flow microcalorimeter. A methodology was developed to characterize rapid and global changes in metabolism and energy flow, not easily detectable with reductionist approaches. Sea water was pumped continuously, 5–10 mm above the sediment, in experimental microcosms; a 100-μm filter prevented passage of meiofauna. This “circulating interface” was taken through the microcalorimeter and from there to an oxygen electrode, and was returned to the microcosm. The microcosms were experimentally eutrophicated using peptone (4 mg·ml ^?1). The relationship between heat production and oxygen tension in the circulating interface has been compared with ATP production, ¹⁴CO₂ and [¹⁴C]particulate matter turnovers. Initial heat steady-state production rises to a peak of 130 to 180 μW·ml^?1 in 6 to 8 h after peptone treatment. The microcalorimetric peak is closely correlated with ¹⁴CO₂ turnover and partially correlated with micro-events on the pO₂ curve. ATP concentration and particulate-¹⁴C turnover increase constantly and then stabilize, with the establishment of a new heat production steady state. The approach provides an indication of the temporal behaviour of complex mixtures of microorganisms and ciliates at the water-sediment interface, and gives holistic measurements of energy flow after induced perturbation (eutrophication) of the ecosystem. Although many problems remain to be solved in this field, it is shown here that flow microcalorimetric measurements can be used to monitor the effects of addition of reagents like pollutants and nutrients.