TPL2 kinase activity regulates microglial inflammatory responses and promotes neurodegeneration in tauopathy mice.

Tumor progression locus 2 (TPL2) (MAP3K8) is a central signaling node in the inflammatory response of peripheral immune cells. We find that TPL2 kinase activity modulates microglial cytokine release and is required for microglia-mediated neuron death in vitro. In acute in vivo neuroinflammation settings, TPL2 kinase activity regulates microglia activation states and brain cytokine levels. In a tauopathy model of chronic neurodegeneration, loss of TPL2 kinase activity reduces neuroinflammation and rescues synapse loss, brain volume loss, and behavioral deficits. Single-cell RNA sequencing analysis indicates that protection in the tauopathy model was associated with reductions in activated microglia subpopulations as well as infiltrating peripheral immune cells. Overall, using various models, we find that TPL2 kinase activity can promote multiple harmful consequences of microglial activation in the brain including cytokine release, iNOS (inducible nitric oxide synthase) induction, astrocyte activation, and immune cell infiltration. Consequently, inhibiting TPL2 kinase activity could represent a potential therapeutic strategy in neurodegenerative conditions.

GABA release by basket cells onto Purkinje cells, in rat cerebellar slices, is directly controlled by presynaptic purinergic receptors, modulating Ca2+ influx.

In many brain regions, Ca(2+) influx through presynaptic P2X receptors influences GABA release from interneurones. In patch-clamp recordings of Purkinje cells (PCs) in rat cerebellar slices, broad spectrum P2 receptor antagonists, PPADS (30microM) or suramin (12microM), result in a decreased amplitude and increased failure rate of minimal evoked GABAergic synaptic currents from basket cells. The effect is mimicked by desensitizing P2X1/3-containing receptors with alpha,beta-methylene ATP. This suggests presynaptic facilitation of GABA release via P2XR-mediated Ca(2+) influx activated by endogenously released ATP. In contrast, activation of P2Y4 receptors (using UTP, 30microM, but not P2Y1 or P2Y6 receptor ligands) results in inhibition of GABA release. Immunological studies reveal the presence of most known P2Rs in >or=20% of GABAergic terminals in the cerebellum. P2X3 receptors and P2Y4 receptors occur in approximately 60% and 50% of GABAergic synaptosomes respectively and are localized presynaptically. Previous studies report that PC output is also influenced by postsynaptic purinergic receptors located on both PCs and interneurones. The high Ca(2+) permeability of the P2X receptor and the ability of ATP to influence intracellular Ca(2+) levels via P2Y receptor-mediated intracellular pathways make ATP the ideal transmitter for the multisite bidirectional modulation of the cerebellar cortical neuronal network.

The unique probe selector: a comprehensive web service for probe design and oligonucleotide arrays.

BACKGROUND: Nucleic acid hybridization, a fundamental technique in molecular biology, can be modified into very effective and sensitive methods for detecting particular targets mixed with millions of non-target sequences. Therefore, avoiding cross-hybridization is the most crucial issue for developing diagnostic methods based on hybridization. RESULTS: To develop a probe with a high discriminating power, this study constructed a web service, the Unique Probe Selector (UPS), for customized probe design. The UPS service integrates a probe design mechanism and a scoring system for evaluating the performance of probe annealing and the uniqueness of a probe in a user-defined genetic background. Starting from an intuitive web interface, the UPS accepts a query with single or multiple sequences in fasta format. The best probe(s) for each sequence can be downloaded from result pages in a fasta or .csv format with a summary of probe characteristics. The option "Unique probe within group" selects the most unique probe for each target sequence with low probability to hybridize to the other sequences in the same submitted query. The option "Unique probe in the specific organism" devises probes for each submitted sequence to identify its target among selected genetic backgrounds based on Unigene. CONCLUSION: The UPS evaluates probe-to-target hybridization under a user-defined condition in silico to ensure high-performance hybridization and minimizes the possibility of non-specific reactions. UPS has been applied to design human arrays for gene expression studies and to develop several small arrays of gene families that were inferred as molecular signatures of cancer typing/staging or pathogen signatures. Notably, UPS is freely accessible at http://array.iis.sinica.edu.tw/ups/.

Serum- and glucocorticoid-inducible kinase 1 enhances zif268 expression through the mediation of SRF and CREB1 associated with spatial memory formation.

Serum- and glucocorticoid-inducible kinase 1 (SGK1) has been shown to play an important role in spatial memory formation, but the molecular mechanism underlying this effect of SGK1 was not known. zif268 is an immediate early gene that is induced by water maze learning. To investigate the role of SGK1 in the regulation of zif268 expression, the dominant negative mutant of SGK1, SGK1 S422A, was infused to the hippocampal CA1 area of rats, and was found to decrease significantly the mRNA level of zif268 in both naïve animals and trained animals. SGK1 was also found to phosphorylate serum response factor (SRF) at Ser73, Ser75, and Ser99, and phosphorylate CREB1 at Ser133. Inhibition of SGK1 phosphorylation sites on SRF and CREB1 with alanine substitution significantly diminished SGK1-enhanced zif268 expression in the promoter-luciferase assay. SGK1 also phosphorylates Elk-1 and SGK1 phosphorylation of Elk-1 decreased the transcriptional activity of Elk-1. But SGK1 phosphorylation of Elk-1 did not affect SGK1-enhanced zif268 expression. Moreover, the phosphorylation of SGK1 was increased in rat CA1 area after water maze learning, accompanied by increased phosphorylation of SRF at Ser99 and increased phosphorylation of CREB1 at Ser133. All these effects were antagonized by SGK1 S422A transfection. These results together suggest that SGK1 enhances zif268 expression through the mediation of SRF and CREB1, and these signaling pathways are associated with spatial memory formation in rats.