The application of lucifferase as a reporter of environmental regulation of gene expression in mycobacteria

This paper describes the construction of pSG10, the first mycobacterial promoter probe shuttle vector to use the structural gene of a bacterial luciferase as a reporter gene. To examine the utility of using bacterial luciferase to measure gene expression in mycobacteria, the authors have used this vector to monitor the induction of the acetamidase gene promoter of Mycobacterium smegmatis. Luciferase proved to be a rapid, sensitive and easily assayable reporter of changes in gene activity in response to environment in mycobacteria.     

New reducing sugar assay for the study of cellulases

A simple assay for reducing sugars based on the production of a coloured formazan and solvent extraction has been developed and used to study the cellulase complex of a Gram-negative bacterium. This assay is more sensitive than others previously described and allows direct study of unconcentrated culture supernatants. The levels of enzyme activity in subcellular fractions were measured after growth on different carbohydrate sources.     

Is a natural ligand of the T lymphocyte CD2 molecule a sulfated carbohydrate?

Recent studies have implicated sulfated polysaccharide (SP) recognition in a range of cell adhesion systems. Inasmuch as the CD2 (E rosette receptor, T11, LFA-2) molecule of human T lymphocytes is a cell surface glycoprotein involved in the adhesion of T cells to various target cells the possibility that CD2 binds SP was investigated. It was found that E rosetting of human T lymphocytes, a phenomenon involving CD2, was readily inhibited by the SP dextran sulfate (DxS) and, to a lesser extent, by the sulfated polymer polyvinyl sulfate whereas 11 other SP had no effect on E rosetting, this effect occurring at the T cell level. mAb binding studies revealed that DxS and polyvinyl sulfate, but none of the other SP tested, inhibited the binding to T cells of the anti-CD2 mAb OKT11 and anti-T112 but augmented expression of the T113 epitope of the CD2 molecule. In contrast, DxS had little or no effect on the binding of anti-CD3, -CD4, -CD8, -Pgp-1 and WT31 (TCR alpha/beta) mAb. Direct evidence that CD2 binds DxS was demonstrated by the ability of DxS-coupled fibers to totally deplete the CD2 Ag from lysates of radiolabeled human T lymphocytes and by the quantitative recovery of the CD2 Ag in fiber eluates. Control fibers coupled with other SP bound little or no CD2. Collectively, the data indicate that the CD2 molecule specifically binds DxS and suggest that a potential target cell ligand for CD2 is a sulfated carbohydrate structure.     

Steroid analogs inhibit hormone binding by an extract from Nippostrongylus brasiliensis (Nematoda)

1. An extract from the rodent nematode Nippostrongylus brasiliensis contained putative receptors that bound radiolabeled sex hormones, based on isoelectric focusing. 2. Binding of radiolabeled testosterone by receptors at pH 4.4 was highly inhibited by the androgen analogs, testosterone-3-oxime and 4-aza-5-androsten-3-on-17 beta-ol. 3. Binding of radiolabeled progesterone by receptors at pH 6.4 was highly inhibited by the progesterone analogs 3,5-seco-4-norpregnan-5-on-3-oic acid and 19-norethisterone or 21-deoxycorticosterone. 4. Binding of radiolabeled 17 beta-estradiol by receptors at pH 4.9 was highly inhibited by epiandrosterone. 5. In vivo development of N. brasiliensis to the adult was partially inhibited by selected steroid analogs.     

Inhibition of in vitro antibody synthesis by cyclophosphamide-induced suppressor cells

A population of suppressor lymphocytes appears in the spleens of mice 5 to 14 days after treatment with a high dose of cyclophosphamide (100–200 mg/kg body wt). Removal of carbonyl iron adherent cells or Ig^? cells from cyclophosphamide (CP)-treated spleen cells does not abolish suppressive activity. These suppressors are, however, sensitive to removal by treatment with anti-Thy-1.2 and rabbit complement. CP-treated spleen cells can suppress the in vitro primary response of normal spleen cells to the soluble hapten-protein conjugate DNP-MON or the particulate antigen HRBC when added at time of culture initiation or up to the second day of culture. CP-treated spleen cells can themselves respond in vitro to DNP-MON, as well as to HRBC, but with altered kinetics from that of normal spleen cells. Collectively, the data suggest that the CP-induced suppressors act late in the in vitro antibody response, possibly by prematurely shutting off antibody synthesis by B cells.     

Effects of thymus-independent (B) cells and the H-2 gene complex on antiviral function of immune thymus-derived (T) cells.

Antiviral activity in vivo exerted by ectromelia virus-immune spleen cells transferred to ectromelia-infected recipients and cytotoxicity against virus-infected target cells in vitro were both properties of non-immunoglobulin (Ig)-bearing cells (which included T cells). Ig-bearing cells, including thymus-independent (B) cells and antibody-secreting cells, were much less active in vivo when injected alone and tended to block rather than amplify the effect triggered by T cells. Ig-bearing cells were also slightly active in vitro, possibly because some T cells have detectable Ig. Antiviral effects in cell transfer experiments were seen only when immune cell donors and infected recipients shared the same H-2 gene complex. These results are consistent with the hypothesis that the T cell response to ectromelia infection is directed against specific virus-induced change(s) in antigen(s), specified by gene(s) in the H-2 complex, which appear in virus-infected cells.