Expression in Bacillus subtilis of the gene for human urogastrone using synthetic ribosome binding sites

Summary A chemically synthesised gene coding for human urogastrone which was earlier cloned in E. coli (Smith et al. 1982) has now been cloned into expression vectors for Bacillus subtilis Two types of constructs have been made, one giving production of methionylurogastrone and the other giving rise to a methionyl-urogastrone- galactosidase fusion polypeptide facilitating quantification of expression levels.The ribosome binding sites used in the expression plasmids are synthetically made oligonucleotides residing on short restriction fragments to allow easy replacement by other ribosome binding sites.Using shuttle vectors and constitutive promoters from Bacillus phages 105 and SPP1, we were able to detect levels of expression amounting to a few thousand molecules per cell during logarithmic growth in both E. coli and B. subtilis.     

Interaction of vasoactive intestinal peptide with mouse lymphocytes: specific binding and the modulation of mitogen responses

Binding of radioiodinated vasoactive intestinal peptide (VIP) to mouse lymphocytes has been investigated. Specific cell binding of 125I-VIP was demonstrated with lymphocytes from mesenteric lymph nodes, subcutaneous lymph nodes, spleen, and Peyer's patches. The binding of VIP by these cells was accounted for by VIP binding sites upon T cells rather than non-T cells. In the presence of VIP, the in vitro response of lymphocytes to the T cell mitogens concanavalin A (Con A) and phytohemagglutinin (PHA) was inhibited in a dose-dependent fashion, whereas that to the B cell mitogen lipopolysaccharide (LPS) was not. There was a close correlation between the potency of VIP and some structurally related peptides for inhibition of 125I-VIP binding and the effect of those peptides on T cell mitogen responses. These observations demonstrate that mouse T lymphocytes have specific VIP receptors and that VIP can modulate the response of T cells to mitogenic stimulation. VIP may be an important immunoregulatory molecule, and may be implicated in the regulation of T cell function in mucosal tissues innervated by VIP-containing neurons.     

Interleukin 2 administered in vivo induces the growth of cultured T cells in vivo

The capacity of exogenous IL 2 to induce the growth of antigen-activated T lymphocytes in vivo was evaluated. The in vivo growth of adoptively transferred T lymphocytes that had been previously cultured long-term with IL 2 was initially examined, because in vitro such T cells are exquisitely dependent upon exogenous IL 2 for proliferation and survival. Daily administration of IL 2 in vivo, beginning on the day of cell transfer, induced these IL 2-dependent long-term cultured T lymphocytes to proliferate in vivo, and the magnitude of in vivo growth was proportional to the dose of IL 2 administered. The capacity of IL 2 to induce the in vivo growth of antigen-activated T cells not previously exposed in vitro to exogenous IL 2 was similarly studied. T lymphocytes from the spleens of immune mice, activated by 5-day culture with tumor antigen before transfer, survived poorly in vivo when injected with antigen alone, but demonstrated marked proliferation in vivo in response to antigen and exogenous IL 2. By contrast, immune spleen cells transferred with antigen, but without prior culture, proliferated without supplementary exogenous IL 2. Moreover, the growth of noncultured donor T cells was not augmented by the administration of exogenous IL 2, implying that noncultured spleen cells immune to tumor antigens can produce sufficient amounts of endogenous IL 2 in vivo to sustain maximal T cell growth over the time period examined. Importantly, the ability of exogenous IL 2 to induce donor T cell growth in vivo correlated with its ability to function in vivo to augment the anti-tumor efficacy of specifically immune donor T cells in models for the adoptive therapy of disseminated antigenic murine leukemia. Thus, the current studies highlight the potential of exogenous IL 2 to induce T cell growth in vivo and suggest that the administration of IL 2 in vivo may be useful for augmenting T cell responses that are relatively deficient in the production of endogenous IL 2.     

The association of medial collagenous tissue with atheroma formation in the aging human aorta as revealed by a special technique

A new technique which brilliantly colors collagen fibers in a field of polarized light reveals that during mid-life the smooth muscle cells in the tunica media of the human aorta begin to disappear. The connective tissue is divided between two regions; one below the subintimal layer and the other under the adventitia. Fine collagen fibers extend upward from the former into the subintima and beyond into the intima and the overlying atheromatous plaques of the aging aorta. Thus, the source of fibrous thickening of the vessel is not confined solely to the intimal layer; at least, a portion of the total collagen content arises deep within the aortic wall.     

DNA-dependent RNA polymerase from Spirochaeta aurantia

Abstract A DNA-dependent RNA polymerase was isolated from Spirochaeta aurantia . The M _r values of the holoenzyme subunits are 164000, 142000, 84000, and 44500. The RNA polymerase activity was sensitive to heparin, streptolydigin, and actinomycin D, while rifampicin and streptovaricin did not inhibit activity.     

Inhibition of Spirochaeta aurantia chemotaxis by neurotoxins.

The effects of neurotoxic compounds on the chemotactic response of Spirochaeta aurantia were investigated. In the presence of neurotoxins that affect action potential generation and transmission in excitable eucaryotic cells, D-xylose taxis was inhibited by 69 to 93%. Inhibition of chemotaxis was not due to decreased viability or motility. This study supports the hypothesis that the molecular basis for sensory signal transduction in S. aurantia involves ion fluxes across the cytoplasmic membrane.     

Apparent Mutagenic Effect of Induction of Lambda Prophage Inserted Between lysA and thyA

Induction of a heat-inducible abnormal lambda prophage inserted between lysA and thyA in Escherichia coli resulted in a number of auxotrophic mutants in the surviving cured-cell populations. These mutants could not be accounted for by deletions arising on formation of lambda hybrid particles carrying regions adjacent to the insertion site. The properties of these mutants, which were almost all spontaneously revertable, have been described and mapped by F′ episome complementation. Tentatively, it was suggested that induction of the lambda lysogen leads to a mutagenic state.     

Side effects of dextroamphetamine therapy of hyperactive children

Dextroamphetamine, prescribed in the treatment of hyperactive children, was associated with significant personality deterioration in five of 26 treated cases. Discontinuance of the drug and, in some cases, substitution of others was followed by lessened symptoms of disorganization or of toxicity. In general, children being treated with psychostimulants should be kept under careful observation for untoward reactions.