Protein kinase A inhibitors enhance radiation-induced apoptosis

In addition to a role for de novo protein synthesis in apoptosis we have previously shown that activation of a protein phosphatase or loss of activity of a kinase is also important in radiation-induced apoptosis in human cells [Baxter, and Lavin (1992): J Immunol 148:149–1954]. We show here that some inhibitors of protein kinases exacerbate radiation-induced apoptosis in the human cell line BM13674. The specific protein kinase A inhibitor isoquinoline sulfonamide (20 μM) gave rise to significantly increased levels of apoptosis at 2–6 h postirradiation compared to values after radiation exposure only. The same concentration of isoquinolinesulfonamide, which was effective in increasing apoptosis, reduced activity markedly. A 66% inhibition of cyclic AMP-dependent protein kinase A activity occurred in unirradiated cells at this concentration of H89 and activity was reduced to 58% in irradiated cells. Calphostin C, a specific inhibitor of protein kinase C, at a concentration of 0.1 μM, which caused 68% inhibition of enzyme activity in irradiated cells, failed to enhance the level of radiation-induced apoptosis. Other kinase inhibitors did not lead to an additional increase in apoptosis over and above that observed after irradiation. The results obtained here provide further support for an important role for modification of existing proteins during radiation-induced apoptosis.     

Outer membrane proteins of Pseudomonas

In this review, we describe the outer membrane proteins of Pseudomonas aeruginosa and related strains from the Pseudomonas fluorescens rRNA homology group of the Pseudomonadaceae, with emphasis on the physiological function and biochemical characteristics of these proteins. The use of opr (for outer membrane protein) is proposed as the genetic designation for the P. aeruginosa outer membrane proteins and letters are assigned, in conjunction with this designation, to known outer membrane proteins. Proteins whose primary functions involve pore formation, transport of specific substrates, cell structure determination and membrane stabilization are discussed. The conservation of selected proteins in the above Pseudomonas species is also examined.     

Association of the type II cAMP-dependent protein kinase with a human thyroid RII-anchoring protein. Cloning and characterization of the RII-binding domain.

The type II cAMP-dependent protein kinase (PKA) is localized to specific subcellular environments through binding of the dimeric regulatory subunit (RII) to anchoring proteins. Subcellular localization is likely to influence which substrates are most accessible to the catalytic subunit upon activation. We have previously shown that the RII-binding domains of four anchoring proteins contain sequences which exhibit a high probability of amphipathic helix formation (Carr, D. W., Stofko-Hahn, R. E., Fraser, I. D. C., Bishop, S. M., Acott, T. E., Brennan, R. G., and Scott J. D. (1991) J. Biol. Chem. 266, 14188-14192). In the present study we describe the cloning of a cDNA which encodes a 1015-amino acid segment of Ht 31. A synthetic peptide (Asp-Leu-Ile-Glu-Glu-Ala-Ala-Ser-Arg-Ile-Val-Asp-Ala-Val-Ile-Glu-Gln-Val -Lys-Ala-Ala-Tyr) representing residues 493-515 encompasses the minimum region of Ht 31 required for RII binding and blocks anchoring protein interaction with RII as detected by band-shift analysis. Structural analysis by circular dichroism suggests that this peptide can adopt an alpha-helical conformation. Both Ht 31 (493-515) peptide and its parent protein bind RII alpha or the type II PKA holoenzyme with high affinity. Equilibrium dialysis was used to calculate dissociation constants of 4.0 and 3.8 nM for Ht 31 peptide interaction with RII alpha and the type II PKA, respectively. A survey of nine different bovine tissues was conducted to identify RII binding proteins. Several bands were detected in each tissues using a 32P-RII overlay method. Addition of 0.4 microM Ht 31 (493-515) peptide to the reaction mixture blocked all RII binding. These data suggest that all anchoring proteins bind RII alpha at the same site as the Ht 31 peptide. The nanomolar affinity constant and the different patterns of RII-anchoring proteins in each tissue suggest that the type II alpha PKA holoenzyme may be specifically targeted to different locations in each type of cell.     

HYBRIDIZATION OF PANUS TIGRINUS AND LENTODIUM SQUAMULOSUM

Crosses between monokaryotic isolates of two strains of Lentodium squamulosum and one strain of Panus tigrinus showed complete intercompatibility. Hybrid fruits from crosses between monokaryotic isolates of L. squamulosum and P. tigrinus were uniformly like the P. tigrinus parent. Though both isolates of L. squamulosum showed considerable variability in form, merging toward P. tigrinus, L. squamulosum could always be distinguished from P. tigrinus from hand sections of the hymenophore. The significance of these results as they bear on the problem of inter-relationships between hymenomycetes and gasteromycetes is discussed. L. squamulosum is recognized as variety squamulosus of P. tigrinus.     

Electron microscopy for the diagnosis of tumours.

The role of leukocyte transfusions in the prevention and treatment of infections in adults with granulocytopenia was investigated. Leukocytes were obtained from healthy volunteers by continuous-flow centrifugation. Histocompatibility antigen (HLA)-matched leukocytes were used to assess the prophylactic value of leukocyte transfusions. Seven patients with acute myelogenous leukemia received HLA-matched leukocytes during the period of maximal granulocytopenia associated with initial remission induction therapy; 20 concurrently treated patients who did not receive leukocyte transfusions were the control group. The patients receiving HLA-matched leukocytes had significantly fewer (P = 0.043) infectious episodes (not bacteriologically proven) during the study period, and remission occurred in 5 of the 7, compared with 10 of the 20 controls. In addition, 52 series of two or more ABO-compatible transfusions were given to 50 patients with proven infection or elevated temperature presumed due to infection and a granulocyte count of less than 0.5 X 10(9)/L. Response, indicated by a decrease in temperature, occurred in 23 patients. Leukocyte transfusions thus have an important adjuvant role in the management of patients with severe granulocytopenia.