Microglial cells are the resident macrophages in the central nervous system. These cells of mesodermal/mesenchymal origin migrate into all regions of the central nervous system, disseminate through the brain parenchyma, and acquire a specific ramified morphological phenotype termed "resting microglia." Recent studies indicate that even in the normal brain, microglia have highly motile processes by which they scan their territorial domains. By a large number of signaling pathways they can communicate with macroglial cells and neurons and with cells of the immune system. Likewise, microglial cells express receptors classically described for brain-specific communication such as neurotransmitter receptors and those first discovered as immune cell-specific such as for cytokines. Microglial cells are considered the most susceptible sensors of brain pathology. Upon any detection of signs for brain lesions or nervous system dysfunction, microglial cells undergo a complex, multistage activation process that converts them into the "activated microglial cell." This cell form has the capacity to release a large number of substances that can act detrimental or beneficial for the surrounding cells. Activated microglial cells can migrate to the site of injury, proliferate, and phagocytose cells and cellular compartments.

Physiology of Microglia

In electrically nonexcitable cells, Ca2+ influx is essential for regulating a host of kinetically distinct processes involving exocytosis, enzyme control, gene regulation, cell growth and prolifera...

Store-operated calcium channels.

Antigen stimulation of immune cells triggers Ca2+ entry through Ca2+ release-activated Ca2+ (CRAC) channels, promoting the immune response to pathogens by activating the transcription factor NFAT. We have previously shown that cells from patients with one form of hereditary severe combined immune deficiency (SCID) syndrome are defective in store-operated Ca2+ entry and CRAC channel function. Here we identify the genetic defect in these patients, using a combination of two unbiased genome-wide approaches: a modified linkage analysis with single-nucleotide polymorphism arrays, and a Drosophila RNA interference screen designed to identify regulators of store-operated Ca2+ entry and NFAT nuclear import. Both approaches converged on a novel protein that we call Orai1, which contains four putative transmembrane segments. The SCID patients are homozygous for a single missense mutation in ORAI1, and expression of wild-type Orai1 in SCID T cells restores store-operated Ca2+ influx and the CRAC current (I(CRAC)). We propose that Orai1 is an essential component or regulator of the CRAC channel complex.

/pdf/a-mutation-in-orai1-causes-immune-deficiency-by-abrogating-4zrmmmi4um.pdf

A mutation in Orai1 causes immune deficiency by abrogating CRAC channel function

Extracellular stimuli elicit changes in gene expression in target cells by activating intracellular protein kinase cascades that phosphorylate transcription factors within the nucleus. One of the best characterized stimulus-induced transcription factors, cyclic AMP response element (CRE)-binding protein (CREB), activates transcription of target genes in response to a diverse array of stimuli, including peptide hormones, growth factors, and neuronal activity, that activate a variety of protein kinases including protein kinase A (PKA), pp90 ribosomal S6 kinase (pp90RSK), and Ca2+/calmodulin-dependent protein kinases (CaMKs)[corrected]. These kinases all phosphorylate CREB at a particular residue, serine 133 (Ser133), and phosphorylation of Ser133 is required for CREB-mediated transcription. Despite this common feature, the mechanism by which CREB activates transcription varies depending on the stimulus. In some cases, signaling pathways target additional sites on CREB or proteins associated with CREB, permitting CREB to regulate distinct programs of gene expression under different conditions of stimulation. This review discusses the molecular mechanisms by which Ser133-phosphorylated CREB activates transcription, intracellular signaling pathways that lead to phosphorylation of CREB at Ser133, and features of each signaling pathway that impart specificity at the level of CREB activation.

CREB: a stimulus-induced transcription factor activated by a diverse array of extracellular signals.

STIM Is a Ca2+ Sensor Essential for Ca2+-Store-Depletion-Triggered Ca2+ Influx

Store-operated Ca2+ (SOC) channels regulate many cellular processes, but the underlying molecular components are not well defined. Using an RNA interference (RNAi)-based screen to identify genes that alter thapsigargin (TG)-dependent Ca2+ entry, we discovered a required and conserved role of Stim in SOC influx. RNAi-mediated knockdown of Stim in Drosophila S2 cells significantly reduced TG-dependent Ca2+ entry. Patch-clamp recording revealed nearly complete suppression of the Drosophila Ca2+ release-activated Ca2+ (CRAC) current that has biophysical characteristics similar to CRAC current in human T cells. Similarly, knockdown of the human homologue STIM1 significantly reduced CRAC channel activity in Jurkat T cells. RNAi-mediated knockdown of STIM1 inhibited TG- or agonist-dependent Ca2+ entry in HEK293 or SH-SY5Y cells. Conversely, overexpression of STIM1 in HEK293 cells modestly enhanced TG-induced Ca2+ entry. We propose that STIM1, a ubiquitously expressed protein that is conserved from Drosophila to mammalian cells, plays an essential role in SOC influx and may be a common component of SOC and CRAC channels.

/pdf/stim1-an-essential-and-conserved-component-of-store-operated-419fhfhgaq.pdf

STIM1, an essential and conserved component of store-operated Ca2+ channel function

Activating transcription factor-1 can mediate Ca(2+)- and cAMP-inducible transcriptional activation.

Ubiquilin 1 Modulates Amyloid Precursor Protein Trafficking and Aβ Secretion

A method of diagnosing a disease with cerebrovascular deposition of amyloid, including Alzheimer's disease, hereditary cerebral hemorrhage with amyloidosis-Dutch type and other amyloidoses, in a mammal is disclosed in which a sample of cerebrospinal fluid is obtained, the level of immunoreactivity toward a monoclonal antibody raised against native PN-2/βAPP or other amyloid precursor protein in the sample is measured, and this measured level is compared to the level of immunoreactivity toward this antibody in a sample from NOTICE OF GOVERNMENT SUPPORT This invention was made with Government support under Grant No. GM-31609 awarded by the National Institutes of Health. The Government has certain rights in this invention. American Cancer Society Grants CD 390 and BC 602/BE 22A provided further support for the development of this invention.

Method of diagnosis of amyloidoses

Immunopurification of Protease Nexin-2 and β amyloid precursor protein is disclosed. Methods of detecting Protease Nexin-2 and the use of these methods in the diagnosis of Alzheimer's disease and other conditions are also disclosed. Additionally, pharmaceutical preparations including Protease Nexin-2 or modified forms thereof are disclosed. Medical uses for the pharmaceutical preparations are also disclosed. These uses include the treatment and prevention of amyloid plaques in Alzheimer's disease and Down's Syndrome.

Steven L. Wagner

Papers

STIM1, an essential and conserved component of store-operated Ca2+ channel function

Activating transcription factor-1 can mediate Ca(2+)- and cAMP-inducible transcriptional activation.

Ubiquilin 1 Modulates Amyloid Precursor Protein Trafficking and Aβ Secretion

Method of diagnosis of amyloidoses

Purification, detection and methods of use of protease nexin-2