Isil Aksan Kurnaz

Ternary complex factor-serum response factor complex-regulated gene activity is required for cellular proliferation and inhibition of apoptotic cell death.

ABSTRACT Members of the ternary complex factor (TCF) subfamily of the ETS-domain transcription factors are activated through phosphorylation by mitogen-activated protein kinases (MAPKs) in response to a variety of mitogenic and stress stimuli. The TCFs bind and activate serum response elements (SREs) in the promoters of target genes in a ternary complex with a second transcription factor, serum response factor (SRF). The association of TCFs with SREs within immediate-early gene promoters is suggestive of a role for the ternary TCF-SRF complex in promoting cell cycle entry and proliferation in response to mitogenic signaling. Here we have investigated the downstream gene regulatory and phenotypic effects of inhibiting the activity of genes regulated by TCFs by expressing a dominantly acting repressive form of the TCF, Elk-1. Inhibition of ternary complex activity leads to the downregulation of several immediate-early genes. Furthermore, blocking TCF-mediated gene expression leads to growth arrest and triggers apoptosis. By using mutant Elk-1 alleles, we demonstrated that these effects are via an SRF-dependent mechanism. The antiapoptotic gene Mcl-1 is identified as a key target for the TCF-SRF complex in this system. Thus, our data confirm a role for TCF-SRF-regulated gene activity in regulating proliferation and provide further evidence to indicate a role in protecting cells from apoptotic cell death.

Astrocyte - neuron lactate shuttle may boost more ATP supply to the neuron under hypoxic conditions - in silico study supported by in vitro expression data

BackgroundNeuro-glial interactions are important for normal functioning of the brain as well as brain energy metabolism. There are two major working models - in the classical view, both neurons and astrocytes can utilize glucose as the energy source through oxidative metabolism, whereas in the astrocyte-neuron lactate shuttle hypothesis (ANLSH) it is the astrocyte which can consume glucose through anaerobic glycolysis to pyruvate and then to lactate, and this lactate is secreted to the extracellular space to be taken up by the neuron for further oxidative degradation.ResultsIn this computational study, we have included hypoxia-induced genetic regulation of these enzymes and transporters, and analyzed whether the ANLSH model can provide an advantage to either cell type in terms of supplying the energy demand. We have based this module on our own experimental analysis of hypoxia-dependent regulation of transcription of key metabolic enzymes. Using this experimentation-supported in silico modeling, we show that under both normoxic and hypoxic conditions in a given time period ANLSH model does indeed provide the neuron with more ATP than in the classical view.ConclusionsAlthough the ANLSH is energetically more favorable for the neuron, it is not the case for the astrocyte in the long term. Considering the fact that astrocytes are more resilient to hypoxia, we would propose that there is likely a switch between the two models, based on the energy demand of the neuron, so as to maintain the survival of the neuron under hypoxic or glucose-and-oxygen-deprived conditions.

ETS-domain transcription factor Elk-1 mediates neuronal survival: SMN as a potential target.

Elk-1 belongs to the ternary complex factors (TCFs) subfamily of the ETS domain proteins, and plays a critical role in the expression of immediate-early genes (IEGs) upon mitogen stimulation and activation of the mitogen-activated protein kinase (MAPK) cascade. The association of TCFs with serum response elements (SREs) on IEG promoters has been widely studied and a role for Elk-1 in promoting cell cycle entry has been determined. However, the presence of the ETS domain transcription factor Elk-1 in axons and dendrites of post-mitotic adult brain neurons has implications for an alternative function for Elk-1 in neurons other than controlling proliferation. In this study, possible alternative roles for Elk-1 in neurons were investigated, and it was demonstrated that blocking TCF-mediated transactivation in neuronal cells leads to apoptosis through a caspase-dependent mechanism. Indeed RNAi-mediated depletion of endogenous Elk-1 results in increased caspase activity. Conversely, overexpression of either Elk-1 or Elk-VP16 fusion proteins was shown to rescue PC12 cells from chemically-induced apoptosis, and that higher levels of endogenous Elk-1 correlated with longer survival of DRGs in culture. It was shown that Elk-1 regulated the Mcl-1 gene expression required for survival, and that RNAi-mediated degradation of endogenous Elk-1 resulted in elimination of the mcl-1 message. We have further identified the survival-of-motor neuron-1 (SMN1) gene as a novel target of Elk-1, and show that the ets motifs in the SMN1 promoter are involved in this regulation.

From Birth Till Death: Neurogenesis, Cell Cycle, and Neurodegeneration

Neurogenesis in the embryo involves many signaling pathways and transcriptional programs and an elaborate orchestration of cell cycle exit in differentiating precursors. However, while the neurons differentiate into a plethora of different subtypes and different identities, they also presume a highly polar structure with a particular morphology of the cytoskeleton, thereby making it almost impossible for any differentiated cell to re‐enter the cell cycle. It has been observed that dysregulated or forced cell cycle reentry is closely linked to neurodegeneration and apoptosis in neurons, most likely through changes in the neurocytoskeleton. However, proliferative cells still exist within the nervous system, and adult neural stem cells (NSCs) have been identified in the Central Nervous System (CNS) in the past decade, raising a great stir in the neuroscience community. NSCs present a new therapeutic potential, and much effort has since gone into understanding the molecular mechanisms driving differentiation of specific neuronal lineages, such as dopaminergic neurons, for use in regenerative medicine, either through transplanted NSCs or manipulation of existing ones. Nevertheless, differentiation and proliferation are two sides of the same coin, just like tumorigenesis and degeneration. Tumor formation may be regarded as a de‐differentiation of tissues, where cell cycle mechanisms are reactivated in differentiated cell types. It is thus important to understand the molecular mechanisms underlying various brain tumors in this perspective. The recent Cancer Stem Cell (CSC) hypothesis also suggests the presence of Brain Tumor Initiating Cells (BTICs) within a tumor population, although the exact origin of these rare and mostly elusive BTICs are yet to be identified. This review attempts to investigate the correlation of neural stem cells/precursors, mature neurons, BTICs and brain tumors with respect to cell cycle regulation and the impact of cell cycle in neurodegeneration. Anat Rec, 292:1953–1961, 2009.

Synthesis and antimycobacterial activity of some phthalimide derivatives

Structurally modified phthalimide derivatives were prepared through condensation of phthalic and tetrafluorophthalic anhydride with selected sulfonamides with variable yields. All compounds were screened for their antimycobacterium activity against Mycobacterium tuberculosis H37Ra (ATCC 25177) using a micro broth dilution technique. The fluorinated derivatives (compounds 2c, 2d, 2f and 2h) had antimycobacterium activity comparable with classical sulfonamide drugs. The minimum inhibitory concentration (MIC) of compounds 2c, 2d, 2f and 2h was greater than that of isoniazid (MIC<0.02 μg/mL) and in vitro activity was greater than that of pyrazinamide, another first line antimycobacterium drug (MIC 50-100 μg/mL). The new compounds could be considered new lead compounds in the treatment of multi-drug resistant tuberculosis.

Both mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 and phosphatidylinositide-3-OH kinase (PI3K)/Akt pathways regulate activation of E-twenty-six (ETS)-like transcription factor 1 (Elk-1) in U138 glioblastoma cells

Epidermal growth factor (EGF) and its receptor (EGFR) have been shown to play a significant role in the pathogenesis of glioblastoma. In our study, the EGFR was stimulated with EGF in human U138 glioblastoma cells. We show that the activated mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 pathway phosphorylated the E twenty-six (ETS)-like transcription factor 1 (Elk-1) mainly at serine 383 residue. Mitogen-activated protein kinase kinase (MEK) 1/2 inhibitor, UO126 and ERK inhibitor II, FR180204 blocked the Elk-1 phosphorylation and activation. The phosphatidylinositide-3-OH kinase (PI3K)/Akt pathway was also involved in the Elk-1 activation. Activation of the Elk-1 led to an increased survival and a proliferative response with the EGF stimulation in the U138 glioblastoma cells. Knocking-down the Elk-1 using an RNA interference technique caused a decrease in survival of the unstimulated U138 glioblastoma cells and also decreased the proliferative response to the EGF stimulation. The Elk-1 transcription factor was important for the survival and proliferation of U138 glioblastoma cells upon the stimulation of EGFR with EGF. The MAPK/ERK1/2 and PI3K/Akt pathways regulated this response via activation of the Elk-1 transcription factor. The Elk-1 may be one of the convergence points for pathways located downstream of EGFR in glioblastoma cells. Utilization of the Elk-1 as a therapeutic target may lead to a novel strategy in treatment of glioblastoma.

Signaling by Neuronal Tyrosine Kinase Receptors: Relevance for Development and Regeneration

Receptor tyrosine kinase activation by binding of neurotrophic factors determines neuronal morphology and identity, migration of neurons to appropriate destinations, and integration into functional neural circuits as well as synapse formation with appropriate targets at the right time and at the right place. This review summarizes the most important aspects of intraneuronal signaling mechanisms and induced gene expression changes that underlie morphological and neurochemical consequences of receptor tyrosine kinase activation in central and peripheral neurons. Anat Rec, 292:1976–1985, 2009.

Elk-1 interacts with neuronal microtubules and relocalizes to the nucleus upon phosphorylation.

ETS domain transcription factor Elk-1 has been primarily studied in the regulation of genes in response to mitogenic stimuli, however the presence of Elk-1 in axonal projections of largely post-mitotic adult hippocampal sections has been reported. This finding has initially led us to a basic question: how is Elk-1 anchored to neuronal projections? To that end, we have investigated the intracellular localization of Elk-1 and its biochemical interactions with neuronal microtubules in model systems. Our results showed co-localization of Elk-1 with microtubules in hippocampal cultures and SH-SY5Y neuroblastoma cell lines, and have further demonstrated that Elk-1 protein can biochemically interact with microtubules in vitro. Analysis of the protein sequence has indicated many putative microtubule binding domains, with the strongest binding prediction in amino acids 314-325, and our results show that Elk-1 can bind to microtubules through most of these regions, but no interaction was observed through the DNA binding domain, where no putative binding motifs were predicted. We further show that upon serum induction, most of the phospho-Elk-1 translocates to the nucleus, which is independent of translation. We propose that Elk-1 is anchored to neuronal microtubules in resting or unstimulated cells, and upon stimulation is phosphorylated, which relocalizes phospho-Elk-1 to the nucleus in neurons.

Exergetic efficiency of ATP production in neuronal glucose metabolism

The exergetic efficiency of ATP production in the brain is assessed using two competing models: 1) the classical model, where both neuron and the astrocyte metabolise glucose; 2) the astrocyte–neuron lactate shuttle hypothesis (ANLSH), where astrocytes metabolise glucose to lactate, then shuttle it to the neuron for aerobic respiration. Exergy analyses showed that ATP production in neuronal mitochondrion is thermodynamically much more efficient than in the cytoplasm. Comparing the Cumulative Degree of Perfection (CDP) for pyruvate production under both models showed that pyruvate production in the cytoplasm causes less exergy loss when lactate is the energy source. Therefore, we predict that the lactate shuttle model is energetically more favourable to the neuron.

Cytoplasmic-to-nuclear volume ratio affects AP-1 complex formation as an indicator of cell cycle responsiveness

Cytoplasmic volume undergoes a series of changes during mitosis in eukaryotes; in turn, signaling events such as osmotic stress can regulate the cytoplasmic volume in cells. In some organisms, increase in cytoplasmic‐to‐nuclear volume ratio was seen to affect the growth potential in cells, however, the mechanistics of such a regulation, if at all present, was unclear. In a computational model, we have constructed a growth factor‐induced signaling pathway leading to AP‐1 heterodimer formation through transcriptional regulation, and analyzed the effects of increasing the cytoplasmic‐to‐nuclear ratio on c‐jun transcription and AP‐1 complex. We have observed that larger cytoplasmic volumes caused both an increase in the final AP‐1 product and a delay in the time of AP‐1 accumulation.