Characterization of T cell hybridomas raised against a glycopeptide containing the tumor-associated T antigen, (betaGal (1-3) alphaGalNAc-O/Ser)

T cell hybridomas were raised against the glycopeptide S⁷² (Core-1) containing the tumor-associated disaccharide Gal (1–3) GalNAc (Core-1) O-linked to serine at position 72 in the mouse hemoglobin derived decapeptide Hb (67–76). All hybridomas recognized the glycopeptide S⁷² (Core-1). Two of the selected hybridomas responded, however, much better to the S⁷² (Tn) glycopeptide containing the monosaccharide GalNAc O-linked to serine. In addition, one hybridoma cross-responded to the glycopeptide T⁷² (Core-1) having a threonine at position 72 instead of a serine. No cross-responses were found to other glycopeptides consisting of the same hemoglobin peptide with different glycans attached or to the unglycosylated peptides. The T cell receptor V and V usage was clearly diverse. The CDR3 regions demonstrated moreover a predominance of small polar amino acid side chains, and three hybridomas contained a common sequence motif. All the sequenced CDR3 regions contained furthermore a conserved proline-glycine motif. In conclusion, immunization with the disaccharide containing glycopeptides S⁷² (Core-1) created a heterogeneous population of glycopeptide specific T cells with the ability of cross-responding toward related glycopeptides.     

betaFSH,betaLH and growth hormone gene expression in blue gourami (Trichogaster trichopterus,Pallas 1770) during spermatogenesis and male sexual behavior

The relationship between gonadal development (histological evidence for spermiogenesis and/or spermatogenesis), sexual behavior (nest-building) and mRNA levels of gonadotropins (betaFSH and betaLH) and growth hormone (GH) in the male pituitary was investigated. Amplification of betaFSH cDNA showed a significantly higher mRNA level in mature males (whether sexually active or not) than in juveniles. However, following PCR amplification of betaLH cDNA, a significantly higher mRNA level was found in the sexually active group compared to the sexually inactive group. These results suggest that FSH may participate in spermatogenesis, whereas LH is more involved in spermiogenesis. The GH mRNA level increased slightly during the maturation process but no significant differences were found between the groups studied.     

In vitro dephosphorylation inhibits the activity of soybean lysine-ketoglutarate reductase in a lysine-regulated manner

In plant seeds, the essential amino acid lysine auto-regulates its own level by modulating the activity of its catabolic enzyme lysine-ketoglutarate reductase via an intracellular signaling cascade, mediated by Ca²⁺ and protein phosphorylation/dephosphorylation. In the present report, it has been further tested whether the activity of soybean lysine-ketoglutarate reductase, as well as that of saccharopine dehydrogenase, the second enzyme in the pathway of lysine catabolism, are modulated by direct phosphorylation of the bifunctional polypeptide containing both of these linked activities. Incubation of purified lysine-ketoglutarate reductase/ saccharopine dehydrogenase with casein kinase II resulted in a significant phosphorylation of the bifunctional enzyme. Moreover, in vitro dephosphorylation of the bifunctional polypeptide with alkaline phosphatase significantly inhibited the activity of lysine-ketoglutarate reductase, but not of its linked enzyme saccharopine dehydrogenase. The inhibitory effect of alkaline phosphatase on lysine-ketoglutarate reductase activity was dramatically stimulated by binding of lysine to the enzyme. Our results suggest that in plant seeds, active lysine-ketoglutarate reductase is a phospho-protein, and that its activity is modulated by opposing actions of protein kinases and phosphatases. Moreover, this modulation is subject to a compound regulation by lysine.     

Calcium-Dependent Transglutaminase of Rat Sympathetic Ganglion in Development and After Nerve Injury

The activity of transglutaminase (TG) was examined in the rat superior cervical ganglion (SCG) during development and after postganglionic nerve crush. During postnatal development the enzyme activity is increased by sevenfold in parallel to protein content of the ganglion and reaches adult levels by day 35 after birth. The endogenous activity (enzyme activity assayed in the absence of the exogenous substrate) during development is transiently elevated with a peak at day 21 postnatal. In the adult ganglion the enzyme specific activity is evenly distributed in all subcellular compartments, but most of it is contained in the cytosol. Within the first hour after axotomy TG activity is rapidly and transiently elevated. The peak value, 80% above control levels, is attained by 30 min postoperative. At this time the activity is increased in all subcellular fractions, but the endogenous activity is selectively increased in the fraction containing nuclei. The enhanced TG activity after axotomy can be prevented by topical treatments with verapamil, an inhibitor of voltage-dependent calcium fluxes across excitable membranes, or with the calcium chelator EGTA. The results show that intracellular TG activity is present in the SCG and that it increases with postnatal growth of the ganglion. After axotomy the enzyme activity is rapidly and transiently increased in the ganglion and this elevation critically depends on calcium fluxes.     

CRP velocity and short-term mortality in ST segment elevation myocardial infarction

Context: There is a known association between C-reactive protein (CRP) levels and adverse outcomes in patients presenting with ST-elevation myocardial infarction (STEMI). The optimal time frame to measure CRP for risk stratification is not known.

Objective: The aim of the current study was to evaluate the relation between the change in CRP velocity (CRPv) and 30-d mortality among STEMI patients.

Material and methods: We included consecutive patients with a diagnosis of STEMI who presented to Tel-Aviv Medical Center between 2008 and 2014 and had their CRP measured with a wide range assay (wr-CRP) at least twice during the 24?h after admission. CRPv was defined as the change in wr-CRP concentration (mg/l) divided by the change in time (in hours) between the two measurements.

Results: The study population comprised of 492 patients, mean age was 62?±?14, 80% were male. CRPv was significantly higher among patients who died within 30 d of admission (1.42?mg/l versus 0.18?mg/l, p?<?0.001). In a multivariate regression model adjusted to multiple confounders, CRPv was independently associated with 30-d mortality (OR 1.39, 95% CI: 1.20–1.62, p?<?0.001).

Conclusion: CRPv might be an independent and rapidly measurable biomarker for short-term mortality in patients presenting with STEMI.     

Double mutants deficient in cytosolic and thylakoid ascorbate peroxidase reveal a complex mode of interaction between reactive oxygen species, plant development, and response to abiotic stresses

Reactive oxygen species (ROS) play a key signaling role in plants and are controlled in cells by a complex network of ROS metabolizing enzymes found in several different cellular compartments. To study how different ROS signals, generated in different cellular compartments, are integrated in cells, we generated a double mutant lacking thylakoid ascorbate peroxidase (tylapx) and cytosolic ascorbate peroxidase1 (apx1). Our analysis suggests that two different signals are generated in plants lacking cytosolic APX1 or tylAPX. The lack of a chloroplastic hydrogen peroxide removal enzyme triggers a specific signal in cells that results in enhanced tolerance to heat stress, whereas the lack of a cytosolic hydrogen peroxide removal enzyme triggers a different signal, which results in stunted growth and enhanced sensitivity to oxidative stress. When the two signals are coactivated in cells (i.e. tylapx/apx1), a new response is detected, suggesting that the integration of the two different signals results in a new signal that manifests in late flowering, low protein oxidation during light stress, and enhanced accumulation of anthocyanins. Our results demonstrate a high degree of plasticity in ROS signaling in Arabidopsis (Arabidopsis thaliana) and suggest the existence of redundant pathways for ROS protection that compensate for the lack of classical ROS removal enzymes such as cytosolic and chloroplastic APXs. Further investigation of the enhanced heat tolerance in plants lacking tylAPX, using mutants deficient in chloroplast-to-nuclei retrograde signaling, suggests the existence of a chloroplast-generated stress signal that enhances basal thermotolerance in plants.