Transfer of IVET Systems to Other Countries: The German Example

AbstractIn some countries the conviction has arisen that their traditional national systems of Initial Vocational Education and Training (IVET) are no longer sufficient for preparing individuals to meet the requirements of the modern world of work referring to a state of the art that is determined by global competition. An easy way to initiate necessary changes appears to be the use of systems that seem to cause economic success. This is obviously the background of bilateral collaboration agreements between the German government and the governments of Greece, Italy, Latvia, Portugal, Slovakia, and Spain (here called “reform countries”) in the IVET-area that shall help the reform countries to change their IVET systems in a way that they better fit to the needs of the labour market. The “reference model” for this kind of collaboration is the German IVET-system. However, there are many conditions for implementation which need to be reflected with regard to a successful “IVET system transfer”. The article at hand will deliver suggestions how to deal with this fact properly, after the German system of IVET and its strengths and weaknesses have been shortly described

Thrombin induces Egr-1 expression in fibroblasts involving elevation of the intracellular Ca2+ concentration, phosphorylation of ERK and activation of ternary complex factor

<p>Abstract</p> <p>Background</p> <p>The serine protease thrombin catalyzes fibrin clot formation by converting fibrinogen into fibrin. Additionally, thrombin stimulation leads to an activation of stimulus-responsive transcription factors in different cell types, indicating that the gene expression pattern is changed in thrombin-stimulated cells. The objective of this study was to analyze the signaling cascade leading to the expression of the zinc finger transcription factor Egr-1 in thrombin-stimulated lung fibroblasts.</p> <p>Results</p> <p>Stimulation of 39M1-81 fibroblasts with thrombin induced a robust and transient biosynthesis of Egr-1. Reporter gene analysis revealed that the newly synthesized Egr-1 was biologically active. The signaling cascade connecting thrombin stimulation with Egr-1 gene expression required elevated levels of cytosolic Ca²⁺, the activation of diacylgycerol-dependent protein kinase C isoenzymes, and the activation of extracellular signal-regulated protein kinase (ERK). Stimulation of the cells with thrombin triggered the phosphorylation of the transcription factor Elk-1. Expression of a dominant-negative mutant of Elk-1 completely prevented Egr-1 expression in stimulated 39M1-81 cells, indicating that Elk-1 or related ternary complex factors connect the intracellular signaling cascade elicited by activation of protease-activated receptors with transcription of the Egr-1 gene. Lentiviral-mediated expression of MAP kinase phosphatase-1, a dual-specific phosphatase that dephosphorylates and inactivates ERK in the nucleus, prevented Elk-1 phosphorylation and Egr-1 biosynthesis in thrombin stimulated 39M1-81 cells, confirming the importance of nuclear ERK and Elk-1 for the upregulation of Egr-1 expression in thrombin-stimulated lung fibroblasts. 39M1-81 cells additionally express M₁muscarinic acetylcholine receptors. A comparison between the signaling cascades induced by thrombin or carbachol showed no differences, except that signal transduction via M₁muscarinic acetylcholine receptors required the transactivation of the EGF receptor, while thrombin signaling did not.</p> <p>Conclusion</p> <p>This study shows that stimulus-transcription coupling in thrombin-treated lung fibroblasts relies on the elevation of the intracellular Ca²⁺-concentration and the activation of PKC and ERK. In the nucleus, ternary complex factors function as key proteins linking the intracellular signaling cascade with enhanced transcription of the Egr-1 gene. This study further shows that the dominant-negative Elk-1 mutant is a valuable tool to study Elk-1-mediated gene transcription.</p

Glucose Homeostasis and Pancreatic Islet Size Are Regulated by the Transcription Factors Elk-1 and Egr-1 and the Protein Phosphatase Calcineurin

Pancreatic β-cells synthesize and secrete insulin. A key feature of diabetes mellitus is the loss of these cells. A decrease in the number of β-cells results in decreased biosynthesis of insulin. Increasing the number of β-cells should restore adequate insulin biosynthesis leading to adequate insulin secretion. Therefore, identifying proteins that regulate the number of β-cells is a high priority in diabetes research. In this review article, we summerize the results of three sophisticated transgenic mouse models showing that the transcription factors Elk-1 and Egr-1 and the Ca2+/calmodulin-regulated protein phosphatase calcineurin control the formation of sufficiently large pancreatic islets. Impairment of the biological activity of Egr-1 and Elk-1 in pancreatic β-cells leads to glucose intolerance and dysregulation of glucose homeostasis, the process that maintains glucose concentration in the blood within a narrow range. Transgenic mice expressing an activated calcineurin mutant also had smaller islets and showed hyperglycemia. Calcineurin induces dephosphorylation of Elk-1 which subsequently impairs Egr-1 biosynthesis and the biological functions of Elk-1 and Egr-1 to regulate islet size and glucose homeostasis

Expression of the C-Terminal Domain of Phospholipase Cβ3 Inhibits Signaling via Gαq-Coupled Receptors and Transient Receptor Potential Channels

Transient receptor potential (TRP) channels are cation channels that play a regulatory role in pain and thermosensation, insulin secretion, and neurotransmission. It has been proposed that activation of TRP channels requires phosphatidylinositol 4,5-bisphosphate, the major substrate for phospholipase C (PLC). We investigated whether inhibition of PLCβ has an impact on TRP channel signaling. A genetic approach was used to avoid off-target effects observed when using a pharmacological PLCβ inhibitor. In this study, we show that expression of PLCβ1ct and PLCβ3ct, truncated forms of PLCβ1 or PLCβ3 that contain the C-terminal membrane binding domains, almost completely blocked the signal transduction of a Gαq-coupled designer receptor, including the phosphorylation of ERK1/2. In contrast, expression of the helix-turn-helix motif (Hα1—Hα2) of the proximal C-terminal domain of PLCβ3 did not affect Gαq-coupled receptor signaling. PLCβ3ct expression impaired signaling of the TRP channels TRPM3 and TRPM8, stimulated with either prognenolone sulfate or icilin. Thus, the C-terminal domain of PLCβ3 interacts with plasma membrane targets, most likely phosphatidylinositol 4,5-bisphosphate, and in this way blocks the biological activation of TRPM3 and TRPM8, which require interaction with this phospholipid. PLCβ thus regulates TRPM3 and TRPM8 channels by masking phosphatidylinositol 4,5-bisphosphate with its C-terminal domain

TRPM3-Induced Gene Transcription Is under Epigenetic Control

Transient receptor potential M3 (TRPM3) cation channels regulate numerous biological functions, including gene transcription. Stimulation of TRPM3 channels with pregnenolone sulfate activates stimulus-responsive transcription factors, which bind to short cognate sequences in the promoters of their target genes. In addition, coregulator proteins are involved that convert the chromatin into a configuration that is permissive for gene transcription. In this study, we determined whether TRPM3-induced gene transcription requires coactivators that change the acetylation pattern of histones. We used compound A485, a specific inhibitor of the histone acetyltransferases CBP and p300. In addition, the role of bromodomain proteins that bind to acetylated lysine residues of histones was analyzed. We used JQ1, an inhibitor of bromodomain and extra terminal domain (BET) family proteins. The results show that both compounds attenuated the activation of AP-1 and CREB-regulated gene transcription following stimulation of TRPM3 channels. Inhibition of CBP/p300 and BET proteins additionally reduced the transcriptional activation potential of the transcription factors c-Fos and Elk-1. Transcriptional upregulation of the interleukin-8 gene was attenuated by A485 and JQ1, indicating that proinflammatory cytokine expression is controlled by CBP/p300 and bromodomain proteins. We conclude that TRPM3-induced signaling involves transcriptional coactivators and acetyl-lysine-bound bromodomain proteins for activating gene transcription

Ca2+ Microdomains, Calcineurin and the Regulation of Gene Transcription

Ca2+ ions function as second messengers regulating many intracellular events, including neurotransmitter release, exocytosis, muscle contraction, metabolism and gene transcription. Cells of a multicellular organism express a variety of cell-surface receptors and channels that trigger an increase of the intracellular Ca2+ concentration upon stimulation. The elevated Ca2+ concentration is not uniformly distributed within the cytoplasm but is organized in subcellular microdomains with high and low concentrations of Ca2+ at different locations in the cell. Ca2+ ions are stored and released by intracellular organelles that change the concentration and distribution of Ca2+ ions. A major function of the rise in intracellular Ca2+ is the change of the genetic expression pattern of the cell via the activation of Ca2+-responsive transcription factors. It has been proposed that Ca2+-responsive transcription factors are differently affected by a rise in cytoplasmic versus nuclear Ca2+. Moreover, it has been suggested that the mode of entry determines whether an influx of Ca2+ leads to the stimulation of gene transcription. A rise in cytoplasmic Ca2+ induces an intracellular signaling cascade, involving the activation of the Ca2+/calmodulin-dependent protein phosphatase calcineurin and various protein kinases (protein kinase C, extracellular signal-regulated protein kinase, Ca2+/calmodulin-dependent protein kinases). In this review article, we discuss the concept of gene regulation via elevated Ca2+ concentration in the cytoplasm and the nucleus, the role of Ca2+ entry and the role of enzymes as signal transducers. We give particular emphasis to the regulation of gene transcription by calcineurin, linking protein dephosphorylation with Ca2+ signaling and gene expression

cAMP response element binding protein (CREB) activates transcription via two distinct genetic elements of the human glucose-6-phosphatase gene

BACKGROUND: The enzyme glucose-6-phosphatase catalyzes the dephosphorylation of glucose-6-phosphatase to glucose, the final step in the gluconeogenic and glycogenolytic pathways. Expression of the glucose-6-phosphatase gene is induced by glucocorticoids and elevated levels of intracellular cAMP. The effect of cAMP in regulating glucose-6-phosphatase gene transcription was corroborated by the identification of two genetic motifs CRE1 and CRE2 in the human and murine glucose-6-phosphatase gene promoter that resemble cAMP response elements (CRE). RESULTS: The cAMP response element is a point of convergence for many extracellular and intracellular signals, including cAMP, calcium, and neurotrophins. The major CRE binding protein CREB, a member of the basic region leucine zipper (bZIP) family of transcription factors, requires phosphorylation to become a biologically active transcriptional activator. Since unphosphorylated CREB is transcriptionally silent simple overexpression studies cannot be performed to test the biological role of CRE-like sequences of the glucose-6-phosphatase gene. The use of a constitutively active CREB2/CREB fusion protein allowed us to uncouple the investigation of target genes of CREB from the variety of signaling pathways that lead to an activation of CREB. Here, we show that this constitutively active CREB2/CREB fusion protein strikingly enhanced reporter gene transcription mediated by either CRE1 or CRE2 derived from the glucose-6-phosphatase gene. Likewise, reporter gene transcription was enhanced following expression of the catalytic subunit of cAMP-dependent protein kinase (PKA) in the nucleus of transfected cells. In contrast, activating transcription factor 2 (ATF2), known to compete with CREB for binding to the canonical CRE sequence 5'-TGACGTCA-3', did not transactivate reporter genes containing CRE1, CRE2, or both CREs derived from the glucose-6-phosphatase gene. CONCLUSIONS: Using a constitutively active CREB2/CREB fusion protein and a mutant of the PKA catalytic subunit that is targeted to the nucleus, we have shown that the glucose-6-phosphatase gene has two distinct genetic elements that function as bona fide CRE. This study further shows that the expression vectors encoding C2/CREB and catalytic subunit of PKA are valuable tools for the study of CREB-mediated gene transcription and the biological functions of CREB

Transient receptor potential melastatin-3 (TRPM3)-induced activation of AP-1 requires Ca 2+ ions and the transcription factors c-Jun, ATF2, and TCF

Abbreviations: AP-1, activator protein-1; bZIP, basic region leucine zipper; MAP kinase, mitogen activated protein kinase; PKC, protein kinase C; SRE, serum response element; SRF, serum response factor; TRE, 12-O-tetradecanoylphorbol-13-acetate (TPA)-responsive element; TRP, transient MOL #95695

Insulin-Responsive Transcription Factors

The hormone insulin executes its function via binding and activating of the insulin receptor, a receptor tyrosine kinase that is mainly expressed in skeletal muscle, adipocytes, liver, pancreatic β-cells, and in some areas of the central nervous system. Stimulation of the insulin receptor activates intracellular signaling cascades involving the enzymes extracellular signal-regulated protein kinase 1/2 (ERK1/2), phosphatidylinositol 3-kinase, protein kinase B/Akt, and phospholipase Cγ as signal transducers. Insulin receptor stimulation is correlated with multiple physiological and biochemical functions, including glucose transport, glucose homeostasis, food intake, proliferation, glycolysis, and lipogenesis. This review article focuses on the activation of gene transcription as a result of insulin receptor stimulation. Signal transducers such as protein kinases or the GLUT4-induced influx of glucose connect insulin receptor stimulation with transcription. We discuss insulin-responsive transcription factors that respond to insulin receptor activation and generate a transcriptional network executing the metabolic functions of insulin. Importantly, insulin receptor stimulation induces transcription of genes encoding essential enzymes of glycolysis and lipogenesis and inhibits genes encoding essential enzymes of gluconeogenesis. Overall, the activation or inhibition of insulin responsive transcription factors is an essential aspect of orchestrating a wide range of insulin-induced changes in the biochemistry and physiology of insulin-responsive tissues