Evidence that lymphokine -activated killer (LAK) cell function is regulated by both serine and tyrosine kinase pathways

Signal transduction pathways operative in lymphokine activated killer (LAK) cells during execution of cytolytic function have never been characterized. Based on ubiquitous involvement of protein phosphorylation in activation of cytolytic mechanisms used by CTL and NK cells, it was hypothesized that changes in protein phosphorylation should occur when LAK encounter tumor targets. It was further hypothesized that protein kinases would regulate LAK-mediated cytotoxicity. Exposure to either SK-Mel-1 (melanoma) or Raji (lymphoma) targets consistently led to increased phosphorylation of two 65-kD LAK proteins pp65a and -b, with isoelectric points (pI) of 5.1 and 5.2 respectively. Increased p65 phosphorylation was initiated between 1 and 5 min after tumor coincubation, occurred on Ser residues, required physical contact between LAK and tumors, correlated with target recognition, and also occurred after crosslinking Fc$\gamma$RIIIA in the absence of tumors. Both pp65a and -b were tentatively identified as phosphorylated forms of the actin-bundling protein L-plastin, based on pI, molecular weight, and cross-reactivity with specific antiserum. The known biochemical properties of L-plastin suggest it may be involved in regulating adhesion of LAK to tumor targets. The protein tyrosine kinase-specific inhibitor Herb A did not block p65 phosphorylation, but blocked LAK killing of multiple tumor targets at a post-binding stage. Greater than 50% inhibition of cytotoxicity was observed after a 2.5-h pretreatment with 0.125 $\mu$g/ml Herb A. Inhibition occurred over a period in pretreatment which LAK were not dependent upon IL-2 for maintenance of killing activity, supporting the conclusion that the drug interfered with mobilization of cytotoxic function. Granule exocytosis measured by BLT-esterase release from LAK occurred after coincubation with tumors, and was inhibited by Herb A LAK cytotoxicity was dependent upon extracellular calcium, suggesting that granule exocytosis rather than Fas ligand was the principal pathway leading to target cell death. The data indicate that protein tyrosine kinases play a pivotal role in LAK cytolytic function by regulating granule exocytosis, and that tumor targets can activate an adhesion dependent Ser kinase pathway in LAK resulting in phosphorylation of L-plastin. ^

Genetic integration of a nuclear -encoded mitochondrial gene with cardiac function

Cardiovascular disease (CVD) is the leading cause of death in the United States. One manifestation of CVD known to increase mortality is an enlarged, or hypertrophic heart. Hypertrophic cardiomyocytes adapt to increased contractile demand at the genetic level with a re-emergence of the fetal gene program and a downregulation of fatty acid oxidation genes with concomitant increased reliance on glucose-based metabolism. To understand the transcriptional regulatory pathways that implement hypertrophic directives we analyzed the upstream promoter region of the muscle specific isoform of the nuclear-encoded mitochondrial gene, carnitine palmitoyltransferase-1β (CPT-1β) in cultured rat neonatal cardiac myocytes. This enzyme catalyzes the rate-limiting step of fatty acid entry into β-oxidation and is downregulated in cardiac hypertrophy and failure, making it an attractive model for the study of hypertrophic gene regulation and metabolic adaptations. We demonstrate that the muscle-enriched transcription factors GATA-4 and SRF synergistically activate CPT-1β; moreover, DNA binding to cognate sites and intact protein structure are required. This mechanism coordinates upregulation of energy generating processes with activation of the energy consuming contractile promoter for cardiac α-actin. We hypothesized that fatty acid or glucose responsive transcription factors may also regulate CPT-1β. Oleate weakly stimulates CPT-1β activity; in contrast, the glucose responsive Upstream Stimulatory Factors (USF) dramatically depresses the CPT-1β reporter. USF regulates CPT-1β through a novel physical interaction with the cofactor PGC-1 and abrogation of MEF2A/PGC-1 synergistic stimulation. In this way, USF can inversely regulate metabolic gene programs and may play a role in the shift of metabolic substrate preference seen in hypertrophy. Failing hearts have elevated expression of the nuclear hormone receptor COUP-TF. We report that COUP-TF significantly suppresses reporter transcription independent of DNA binding and specific interactions with GATA-4, Nkx2.5 or USF. In summary, CPT-1β transcriptional regulation integrates mitochondrial gene expression with two essential cardiac functions: contraction and metabolic substrate oxidation. ^

Physical, biological and chemical characteristics of Canadian and Danish lakes during summer

1. Winter temperatures differ markedly on the Canadian prairies compared with Denmark. Between 1 January 1998 and 31 December 2002, average weekly and monthly temperatures did not drop below 0 °C in the vicinity of Silkeborg, Denmark. Over this same time, weekly average temperatures near Calgary, Alberta, Canada, often dropped below -10 °C for 3-5 weeks and the average monthly temperature was below 0 °C for 2-4 months. Accordingly, winter ice conditions in shallow lakes in Canada and Denmark differed considerably. 2. To assess the implications of winter climate for lake biotic structure and function we compared a number of variables that describe the chemistry and biology of shallow Canadian and Danish lakes that had been chosen to have similar morphometries. 3. The Danish lakes had a fourfold higher ratio of chlorophyll-a: total phosphorus (TP). Zooplankton : phytoplankton carbon was related to TP and fish abundance in Danish lakes but not in Canadian lakes. There was no significant difference in the ratio log total zooplankton biomass : log TP and the Canadian lakes had a significantly higher proportion of cladocerans that were Daphnia. These differences correspond well with the fact that the Danish lakes have more abundant and diverse fish communities than the Canadian lakes. 4. Our results suggest that severe Canadian winters lead to anoxia under ice and more depauperate fish communities, and stronger zooplankton control on phytoplankton in shallow prairie lakes compared with shallow Danish lakes. If climate change leads to warmer winters and a shorter duration of ice cover, we predict that shallow Canadian prairie lakes will experience increased survivorship of planktivores and stronger control of zooplankton. This, in turn, might decrease zooplankton control on phytoplankton, leading to 'greener' lakes on the Canadian prairies.

Physical oceanography during five ECOMARGE cruises

The first experiment of the ECOMARGE programme (ECOsystèmes de MARGE continentale) was initiated in 1983-1984, in the Gulf of Lions (northwestern Mediterranean Sea). The objectives of the ECOMARGE-I experiment were: to quantify the transfer of particulate matter, in general, and of organic carbon, in particular, from its introduction to and formation in the waters of the continental shelf-to its consumption or sedimentation on the shelf or its transfer to the slope and deep sea; and to understand the processes involved in that transfer, consumption and sedimentation together with their variability in space and time. The results of that experiment, from 1983 to 1988, are presented in this Special Issue. The highlights of the results are summarised in this paper. These results indicate that, of the particles formed in the waters of the continental shelf and those introduced by rivers, some are deposited as sediments on the shelf. A portion is transported offshore, however, to the slope and deep sea. The Rho^ne River, in the northeastern part of the study area, is the major source of continental material; this is transported to sea in a benthic nepheloid layer and, mostly, alongshore to the southwest. Here, it largely leaves the shelf through the canyons, especially the Lacaze-Duthiers Canyon. In the offshore waters, particle concentrations and distributions show surficial, intermediate and benthic nepheloid layers. These turbid structures increase towards the southwest, corresponding to the seaward shift of the front between the coastal waters and the Liguro-Provençal cyclonic gyre, a major forcing function in the Gulf of Lions. Considering the source and fate of particles (largely biogenic from the euphotic zone and abiogenic from deeper waters) a layered system is described, which is emphasized by the concentrations of natural and artificial elements and compounds. Of the flux of particles to the Lacaze-Duthiers Canyon, on a decadal scale, about 30% (as a minimum) is estimated to be stored as sediment; the remainder is transported down-canyon, towards the deep sea. The temporal variability of processes affecting this net seaward transport, of both biogenic and abiogenic material, is from hours, days to seasonal, and probably interannual, time scales. The response of the system to these variations is rapid, with pulses of increased discharge of particles from the adjacent shelf being detected in sediment traps in the Lacaze-Duthiers Canyon in less than 16 days (the temporal resolution of the traps). Based upon the study of tracers of particulate matter and environmental factors (i.e. river discharge and climatic conditions), it appears that the contribution from the Rho^ne River and its adjacent area is maximal during the winter; at this time, the flow of the Liguro-Provençal Current also increases. In contrast, the maximum relative contribution of the adjacent southwesterly area to the flux in the Lacaze-Duthiers Canyon occurs in summer, during storm events.

Physical properties, velocity and attenuation characteristics, and mineralogy of gabbros from ODP Hole 176-735B

Laboratory compressional wave (Vp) and shear wave (Vs) velocities were measured as a function of confining pressure for the gabbros from Hole 735B and compared to results from Leg 118. The upper 500 m of the hole has a Vp mean value of 6895 m/s measured at 200 MPa, and at 500 meters below seafloor (mbsf), Vp measurements show a mean value of 7036 m/s. Vs mean values in the same intervals are 3840 m/s and 3857 m/s, respectively. The mean Vp and Vs values obtained from log data in the upper 600 m are 6520 and 3518 m/s, respectively. These results show a general increase in velocity with depth and the velocity gradients estimate an upper mantle depth of 3.32 km. This value agrees with previous work based on dredged samples and inversion of rare element concentrations in basalts dredged from the conjugate site to the north of the Atlantis Bank. Laboratory measurements show Vp anisotropy ranging between 0.4% and 8.8%, with the majority of the samples having values less than 3.8%. Measurements of velocity anisotropy seem to be associated with zones of high crystal-plastic deformation with predominant preferred mineral orientations of plagioclase, amphiboles, and pyroxenes. These findings are consistent with results on gabbros from the Hess Deep area and suggest that plastic deformation may play an important role in the seismic properties of the lower oceanic crust. In contrast to ophiolite studies, many of the olivine gabbros show a small degree of anisotropy. Log derived Vs anisotropy shows an average of 5.8% for the upper 600 m of Hole 735B and tends to decrease with depth where the overburden pressure and the age of the crustal section suggests closure of cracks and infilling of fractures by alteration minerals. Overall the results indicate that the average shear wave splitting in Hole 735B might be influenced by preferred structural orientations and the average value of shear wave splitting may not be a maximum because structural dips are <90°. The maximum fast-wave orientation values could be influenced by structural features striking slightly oblique to this orientation or by near-field stress concentrations. However, flexural wave dispersion analyses have not been performed to confirm this hypothesis or to indicate to what extent the near-field stresses may be influencing shear wave propagation. Acoustic impedance contrasts calculated from laboratory and logging data were used to generate synthetic seismograms that aid in the interpretation of reflection profiles. Several prominent reflections produced by these calculations suggest that Fe-Ti oxides and shear zones may contribute to the reflective nature of the lower oceanic crust. Laboratory velocity attenuation (Q) measurements from below 500 m have a mean value of 35.1, which is consistent with previous vertical seismic profile (VSP) and laboratory measurements on the upper 500 m.