Identification and functional characterization of novel phosphotyrosine and phosphoserine residues in the interleukin-2 receptor complex

Interleukin-2 (IL-2) is a major T cell growth factor and plays an essential role in the development of normal immune responses. The Janus kinases (Jaks) and Signal transducers and activators of transcription (Stats) are critical for transducing signals from the IL-2 receptors (IL2Rs) to the nucleus to control cell growth and differentiation. In recent years there has been increasing evidence to indicate that the IL-2 activated Jak3/Stat5 pathway provides a new molecular target for immune suppression. Thus, understanding the regulation of this effector cascade has important therapeutic potential.^ One objective of this work was to identify and define the role and molecular mechanism of novel phosphorylation sites in Jak3. Using functional proteomics, three novel Jak3 phosphorylation sites, Y904, Y939 and S574 were identified. Phosphospecific antibodies confirmed that phosphorylation of Y904 and Y939 were mediated by IL-2 and other IL-2 family cytokines in distinct cell types. Biochemical analysis demonstrated that phosphorylation of both Y904 and Y939 positively regulated Jak3 enzymatic activity, while phosphorylation of S574 did not affect Jak3 in vitro kinase activity. However, a gain-of-function mutation of S574 in Jak3 abrogated IL-2 mediated Stat5 activation, suggesting that phosphorylation of this residue might serve a negative role to attenuate IL-2 signaling. Furthermore, mechanistic analysis suggested that phosphorylation of Y904 in Jak3 affects the KmATP of Jak3, while phosphorylation of Y939 in Jak3 was required to bind one of its substrates, Stat5.^ The second objective was to determine the role of serine/threonine phosphatases in the regulation of the IL2R complex. Activation of Jak3 and Stat5 by IL-2 is a transient event mediated by phosphorylation. Using a specific PP1/PP2A inhibitor, we observed that inhibition of PP1/PP2A negatively regulated the IL-2 activated Jak3/Stat5 signaling pathway in a human NK cell line (YT) and primary human T cells. More importantly, coimmunoprecipitation assays indicated that inhibition of PP1/PP2A blocked the formation of an active IL2R complex. Pretreatment of cells with the inhibitor also reduced the electrophoretic mobility of the IL2Rβ and IL2Rγ subunits in YT cells, suggesting that inhibition of PP1/PP2A directly or indirectly regulates undefined serine/threonine kinases which phosphorylate these proteins. Based on these observations, a model has emerged that serine/threonine phosphorylation of the IL2Rβ and IL2Rγ subunits causes a conformational change of these proteins, which disrupts IL2R dimerization and association of Jak3 and Stat5 to these receptors.^

A standard operation procedure and a pilot water testing for herbicides in northwest Harris County, Texas

Herbicides are used to control the growth of weeds along highways, power lines, and many other urban locations. Exposure to herbicides has been linked to adverse health outcomes. This study was initiated to pretest for the presence of herbicides in multiple water sources near intersections in a corridor in the Northwest Harris County (specifically in the Highway 6/FM 1960, North Freeway 45, US 290 and S 99 corridor). Roadside water and tap water samples were collected and analyzed for herbicides using the established Environmental Protection Agency (EPA) Method 515.4: "Determination of Chlorinated Acids in Drinking Water by Liquid-Liquid Micro-extraction, Derivatization, and Fast Gas Chromatography with Electron Capture Detection." A standard operating procedure (adapted from the US EPA Method 515.4) was developed for subsequent, larger studies of environmental fate of herbicides and non-occupational exposure risks. Preliminary testing of 16 water samples was performed to pretest the existence of trace herbicides; all concentrations that were greater than the minimum reporting limits of each analyte are reported with a 99 percent confidence. This study failed to find concentrations above the limits of detection of the method in any of the samples collected on June 15, 2008. However, this does not indicate that the waters around the NW Harris County are free of herbicides and metabolites. A larger and repeated sampling in the region would be necessary to make that claim. ^

Evaluation of Fungicides and Herbicides for Management of White Mold of Soybean

Several new fungicide products are either available or will be available for management of white mold of soybean. This study was conducted at the Muscatine Island Research and Demonstration Farm, and one farmer’s field in northeast Iowa.

Insoluble residues of the fine-grained sediments from DSDP Leg 67 Holes

Insoluble residues of Late Cretaceous to Quaternary deep-sea samples from slope, trench, and oceanic plate sites south of Guatemala were examined, specifically for the distribution of clay minerals in the <2-µm fraction and of silt grains in the 20-63-µm fraction. Widespread "oceanic" particles (biogenic opal, rhyolitic glass) and their diagenetic products (smectite, clinoptilolite, heulandite) were distinguished from terrigenous material - illite, kaolinite, chlorite, plagioclase, quartz, and heavy minerals. The main results of this investigation are: (1) At Site 494 on the slope immediately adjacent to the trench, terrigenous supplies testify to a slope position of the whole sequence back to the Late Cretaceous. (2) At Site 495 on the Pacific Cocos Plate, "oceanic" and terrigenous sedimentation are clearly separated. Whereas the pelagic sedimentation prevailed in the early Miocene, terrigenous minerals appeared in the middle Miocene in the clay fraction, and in the early Pliocene in the coarse silt fraction. These terrigenous supplies are interpreted as having been transported by suspension clouds crossing the slope and even the trench. The alternative, however, an eolian transport, cannot be excluded.

Chemical and Sr isotopic compositions of 3 step leachates and residues of bulk samples, zeolites, and smectites from Lower Miocene sediments at ODP Site 166-1007

Massive clinoptilolite authigenesis was observed at about 1105 meters below sea floor (mbsf) in lower Miocene wellcompacted carbonate periplatform sediments from the Great Bahama Bank [Ocean Drilling Program, ODP Leg 166, Site 1007]. The diagenetic assemblage comprises abundant zeolite crystallized within foraminifer tests and sedimentary matrix, as well as Mg smectites. In carbonate-rich deposits, the formation of the zeolite requires a supply of silica. Thus, the objective of the study is to determine the origin of the silica supply, its diagenetic evolution, and consequently the related implications on interpretation of the sedimentary record, in terms of local or global paleoceanographic change. For lack of evidence for any volcaniclastic input or traces of Si-enriched deep fluids circulation, an in situ biogenic source of silica is validated by isotopic data and chemical modeling for the formation of such secondary minerals in shallow-water carbonate sequences. Geochemical and strontium isotopic data clearly establish the marine signature of the diagenetic zeolite, as well as its contemporaneous formation with the carbonate deposition (Sr model ages of 19.6-23.2 Ma). The test of saturation for the pore fluids specifies the equilibrium state of the present mineralogical assemblage. Seawater-rock modeling specifies that clinoptilolite precipitates from the dissolution of biogenic silica, which reacts with clay minerals. The amount of silica (opal-A) involved in the reaction has to be significant enough, at least 10 wt.%, to account for the observed content of clinoptilolite occurring at the most zeolite-rich level. Modeling also shows that the observed amount of clinoptilolite (~19%) reflects an in situ and short-term reaction due to the high reactivity of primary biogenic silica (opal-A) until its complete depletion. The episodic occurrence of these well-lithified zeolite-rich levels is consistent with the occurrence of seismic reflectors, particularly the P2 seismic sequence boundary located at 1115 mbsf depth and dated as 23.2 Ma. The age range of most zeolitic sedimentary levels (biostratigraphic ages of 21.5-22 Ma) correlates well with that of the early Miocene glaciation Mi-1 and Mi-1a global events. Thus, the clinoptilolite occurrence in the shallow carbonate platform environment far from volcanogenic supply, or in other sensitive marine areas, is potentially a significant new proxy for paleoproductivity and oceanic global events, such as the Miocene events, which are usually recognized in deep-sea pelagic sediments and high latitude deposits.