Drosophila insulin degrading enzyme and rat skeletal muscle insulin protease cleave insulin at similar sites

Insulin degradation is an integral part of the cellular action of insulin. Recent evidence suggests that the enzyme insulin protease is involved in the degradation of insulin in mammalian tissues. Drosophila, which has insulin-like hormones and insulin receptor homologues, also expresses an insulin degrading enzyme with properties that are very similar to those of mammalian insulin protease. In the present study, the insulin cleavage products generated by the Drosophila insulin degrading enzyme were identified and compared with the products generated by the mammalian insulin protease. Both purified enzymes were incubated with porcine insulin specifically labeled with 125I on either the A19 or B26 position, and the degradation products were analyzed by HPLC before and after sulfitolysis. Isolation and sequencing of the cleavage products indicated that both enzymes cleave the A chain of intact insulin at identical sites between residues A13 and A14 and A14 and A15. Sequencing of the B chain fragments demonstrated that the Drosophila enzyme cleaves the B chain of insulin at four sites between residues B10 and B11, B14 and B15, B16 and B17, and B25 and B26. These cleavage sites correspond to four of the seven cleavage sites generated by the mammalian insulin protease. These results demonstrate that all the insulin cleavage sites generated by the Drosophila insulin degrading enzyme are shared in common with the mammalian insulin protease. These data support the hypothesis that there is evolutionary conservation of the insulin degrading enzyme and further suggest that this enzyme plays an important role in cellular function.     

Effects of chloramphenicol on RNA synthesis in Spirodela chlorplasts

RNA from chloroplasts isolated from Spirodela oligorrhiza includedrelatively rapidly-labeled fractions with apparent molecularweights of 2.7; 1.2; 0.7; and 0.5x10⁶. With longer labeling,radioactivity appeared in the mature rRNAs (1.1 and 0.56x10⁶MW). Chloramphenicol inhibited the appearance of labeled maturerRNA, but increased the net labeling and caused the accumulationof the pulse-labeled RNAs, effects similar to those reportedfor bacteria. ¹ 1 Permanent address: Dept. of Biological Sciences, S.U.N.Y.,Binghamton, N.Y. 13901, U.S.A. Supported by a SUNY/ResearchFoundation Faculty Research Fellowship. (Received November 12, 1974; )     

Control of estrogen and androgen receptors in the rat pineal gland by catecholamine transmitter

Sucrose gradient studies of rat pineal cytosol incubated with ³H-estradiol (female pineals) or ³H-5 α -dihydrotestosterone (male pineals) revealed a radioactivity peak in the 8 S region which disappeared after superior cervical ganglionectomy or incubation with excess unlabeled hormone. Ganglionectomy decreased significantly estradiol and testosterone uptake by the pineal gland $\text{in vitro}$ as well as high affinity binding to pineal cytoplasmic and nuclear components. Norepinephrine treatment counteracted all the effects of ganglionectomy but was unable to modify hormone uptake and binding by the pineal gland of sham-operated controls. Pre-treatment with actinomycin D or propranolol but not with phentolamine impaired norepinephrine effects; propranolol blockage however was only partial. Administration of isoproterenol, L-dopa or phentolamine increased hormone uptake by denervated pineals. The effects of isoproterenol were also observed $\text{in vitro}$ and were blocked by propranolol. These results indicate that sex steroid receptors in the pinealocytes are controlled by norepinephrine via beta-adrenergic receptors and that depletion of neural norepinephrine enhanced responsiveness of pineal hormone receptors to exogenous catecholamines.     

Effect of Adenosine and Deoxyadenosine on the Incorporation and Breakdown of Thymidine in Escherichia coli K-12

Deoxyadenosine (AdR) and adenosine (AR) enhance the incorporation of thymidine (TdR) into bacterial deoxyribonucleic acid (DNA) by the inhibition of TdR phosphorolysis in vivo. Neither of the purine nucleosides has an effect on the reaction catalyzed by TdR phosphorylase in vitro. AdR induces TdR phosphorylase and both purine nucleosides induce purine nucleoside phosphorylase. AR can stimulate uptake of more TdR into bacterial DNA than AdR.     

Trichromatic visual system in an insect and its sensitivity control by blue light

Summary The spectral sensitivity of the visual cells in the compound eye of the mothDeilephila elpenor was determined by electrophysiological mass recordings during exposure to monochromatic adapting light. Three types of receptors were identified. The receptors are maximally sensitive at about 350 nm (ultraviolet), 450 nm (violet), and 525 nm (green). The spectral sensitivity of the green receptors is identical to a nomogram for a rhodopsin with _max at 525 nm. The spectral sensitivity of the other two receptors rather well agrees with nomograms for corresponding rhodopsins. The recordings indicate that the green receptors occur in larger number than the other receptors. The ultra-violet and violet receptors probably occur in about equal number.The sensitivity after monochromatic adapting illumination varies with the wavelength of the adapting light, but is not proportional to the spectral sensitivity of the receptors. The sensitivity is proportional to the concentration of visual pigment at photoequilibrium. The equilibrium is determined by the absorbance coefficients of the visual pigment and its photoproduct at each wavelength. The concentration of the visual pigment, and thereby the sensitivity, is maximal at about 450 nm, and minimal at wavelengths exceeding about 570 nm.The light from a clear sky keeps the relative concentration of visual pigment in the green receptors, and the relative sensitivity, at about 0.62. The pigment concentration in the ultra-violet receptors is about 0.8 to 0.9, and that in the violet receptors probably about 0.6. At low ambient light intensities a chemical regeneration of the visual pigments may cause an increase in sensitivity. At higher intensities the concentrations of the visual pigments remain constant. Due to the constant pigment concentrations the input signals from the receptors to the central nervous system contain unequivocal information about variations in intensity and spectral distribution of the stimulating light.The work reported in this article was supported by the Swedish Medical Research Council (grant no B 73-04X-104-02B), by Karolinska Institutet, and by a grant (to G. Höglund) from Deutscher Akademischer Austauschdienst, and by the Deutsche Forschungsgemeinschaft, Schwerpunktsprogramm Rezeptorphysiologie HA 258-10, and SFB 114.     

Phosphorolysis of 5-Fluoro-2′-Deoxyuridine in Escherichia coli and Its Inhibition by Nucleosides

The effect of ribonucleosides and deoxyribonucleosides on the phosphorolysis of 5-fluoro-2'-deoxyuridine (FUdR) was examined in cells of Escherichia coli. All nucleosides tested except guanosine and deoxyguanosine inhibited phosphorolysis. By using genetically marked strains it was found that in vivo FUdR is a specific substrate of thymidine phosphorylase.     

Role of NK cells in protection of mice against herpes simplex virus-1 infection

Natural killer (NK) cells have been implicated in the recognition and killing of a variety of virus infected target cells in vitro, yet their role in vivo remains uncertain. In these experiments, the role of NK cells in the regulation of resistance to herpes simplex virus-1 (HSV-1) was studied. Adult C57BL/6 mice are resistant to HSV-1 (HFEM strain), but are rendered highly susceptible by treatment with cyclophosphamide 24 hr prior to infection. In this model, passive transfer of 10(8) normal spleen cells or 10(7) poly I:C-treated spleen cells provided protection for 72% of the recipients. Spleen cells from NK cell-deficient beige mice similarly treated failed to engender passive protection. The phenotype of the cells responsible for transferring protection was NK1.1+, and asialo GM1+. Transfer of NK cells resulted in marked reduction of HSV titers in the livers and brains of recipients. These experiments provide direct evidence for a role for NK cells in protection against development of fatal HSV infection in mice.     

Detection of chemicals that stimulate Tn9 transposition in Escherichia coli K12

Summary A spot test has been developed for detecting substances that enhance the transposition of Tn9 in Escherichia coli. Phage :: Tn9-infected cells were plated on chloramphenicol media and a drop of the test substance was placed at the center of the plate. Following incubation, chloramphenicol-resistant colonies appeared due to the transposition of Tn9 to the bacterial chromosome. By comparing the test plate and a control plate with respect to the number and distribution of colonies, the effect of the test compound can be evaluated.Out of over 100 compounds tested, acetate, two detergents (Brij 58 and Nonidet P40) and dimethylsulfoxide were found to enhance transposition 3–20 fold. Acetate was also found to enhance the transposition of Tn5 and Tn10. The stimulating effect of Brij 58 was lost when palmitic acid was added with the Brij 58. The nature of these substances, which we refer to as transposagens, suggests an involvement of lipid or membrane in the transposition process.Abbreviations AMP-R, CAM-R, KAN-R, SPC-R, TET-R resistance to ampicillin, chloramphenicol, kanamycin, spectinomycin and tetracycline, respectively - DMSO dimethylsulfoxide A preliminary report of this work was presented at the Fifth Mid-Atlantic Extrachromosomal Genetic Elements Meeting, 1981 (Datta, Randolph and Rosner, Plasmid 7:99, 1982)