The rockefeller university

The autonomic nervous system regulates fuel availability and energy storage in the liver, adipose tissue, and other organs; however, the molecular components of this neural circuit are poorly understood. We sought to identify neural populations that project from the CNS indirectly through multisynaptic pathways to liver and epididymal white fat in mice using pseudorabies virus strains expressing different reporters together with BAC transgenesis and immunohistochemistry. Neurons common to both circuits were identified in subpopulations of the paraventricular nucleus of the hypothalamus (PVH) by double labeling with markers expressed in viruses injected in both sites. The lateral hypothalamus and arcuate nucleus of the hypothalamus and brainstem regions (nucleus of the solitary tract and A5 region) also project to both tissues but are labeled at later times. Connections from these same sites to the PVH were evident after direct injection of virus into the PVH, suggesting that these regions lie upstream of the PVH in a common pathway to liver and adipose tissue (two metabolically active organs). These common populations of brainstem and hypothalamic neurons express neuropeptide Y and proopiomelanocortin in the arcuate nucleus, melanin-concentrating hormone, and orexin in the lateral hypothalamus and in the corticotrophin-releasing hormone and oxytocin in the PVH. The delineation of this circuitry will facilitate a functional analysis of the possible role of these potential command-like neurons to modulate autonomic outflow and coordinate metabolic responses in liver and adipose tissue.

The concentration of the second messenger cAMP is tightly controlled in cells by the activity of phosphodiesterases. We have previously described how the protein kinase A-anchoring protein mAKAP serves as a scaffold for the cAMP-dependent protein kinase PKA and the cAMP-specific phosphodiesterase PDE4D3 in cardiac myocytes. PKA and PDE4D3 constitute a negative feedback loop whereby PKA-catalyzed phosphorylation and activation of PDE4D3 attenuate local cAMP levels. We now show that protein phosphatase 2A (PP2A) associated with mAKAP complexes is responsible for reversing the activation of PDE4D3 by catalyzing the dephosphorylation of PDE4D3 serine residue 54. Mapping studies reveal that a C-terminal mAKAP domain (residues 2085-2319) binds PP2A. Binding to mAKAP is required for PP2A function, such that deletion of the C-terminal domain enhances both base-line and forskolin-stimulated PDE4D3 activity. Interestingly, PP2A holoenzyme associated with mAKAP complexes in the heart contains the PP2A targeting subunit B56 delta. Like PDE4D3, B56 delta is a PKA substrate, and PKA phosphorylation of mAKAP-bound B56 delta enhances phosphatase activity 2-fold in the complex. Accordingly, expression of a B56 delta mutant that cannot be phosphorylated by PKA results in increased PDE4D3 phosphorylation. Taken together, our findings demonstrate that PP2A associated with mAKAP complexes promotes PDE4D3 dephosphorylation, serving both to inhibit PDE4D3 in unstimulated cells and also to mediate a cAMP-induced positive feedback loop following adenylyl cyclase activation and B56 delta phosphorylation. In general, PKA.PP2A.mAKAP complexes exemplify how protein kinases and phosphatases may participate in molecular signaling complexes to dynamically regulate localized intracellular signaling.

Amyloid beta (A beta) is involved in the etiology of Alzheimer's disease (AD) and may contribute to cognitive deficits by increasing internalization of ionotropic glutamate receptors. Striatal-enriched protein tyrosine phosphatase 61 (STEP61), which is targeted in part to the postsynaptic terminal, has been implicated in this process. Here we show that STEP61 levels are progressively increased in the cortex of Tg2576 mice over the first year, as well as in prefrontal cortex of human AD brains. The increased STEP61 was associated with greater STEP activity, dephosphorylation of phospho-tyr(1472) of the NR2B subunit, and decreased NR1 and NR2B subunits on neuronal membranes. Treatment with A beta-enriched medium also increased STEP61 levels and decreased NR1/NR2B abundance in mouse cortical cultures as determined by biotinylation experiments. In STEP knock-out cultures, A beta treatment failed to induce NMDA receptor internalization. The mechanism for the increase in STEP61 levels appears to involve the ubiquitin proteasome system. Blocking the proteasome resulted in elevated levels of STEP61. Moreover, STEP61-ubiquitin conjugates were increased in wild-type cortical slices upon A beta treatment as well as in 12 month Tg2576 cortex. These findings reveal a novel mechanism by which A beta-mediated accumulation of STEP61 results in increased internalization of NR1/NR2B receptor that may contribute to the cognitive deficits in AD.

Many promoters in eukaryotes have nucleosome-depleted regions (NDRs) containing transcription factor binding sites. However, the functional significance of NDRs is not well understood. Here, we examine NDR function in two cell cycle-regulated promoters, CLN2pr and HOpr, by varying nucleosomal coverage of the binding sites of their activator, Swi4/Swi6 cell-cycle box (SCB)-binding factor (SBF), and probing the corresponding transcriptional activity in individual cells with time-lapse microscopy. Nucleosome-embedded SCBs do not significantly alter peak expression levels. Instead, they induce bimodal, "on/off" activation in individual cell cycles, which displays short-term memory, or epigenetic inheritance, from the mother cycle. In striking contrast, the same SCBs localized in NDR lead to highly reliable activation, once in every cell cycle. We further demonstrate that the high variability in Cln2p expression induced by the nucleosomal SCBs reduces cell fitness. Therefore, we propose that the NDR function in limiting stochasticity in gene expression promotes the ubiquity and conservation of promoter NDR.

Entropy rate quantifies the change of information of a stochastic process (Cover & Thomas, 2006). For decades, the temporal dynamics of spike trains generated by neurons has been studied as a stochastic process (Barbieri, Quirk, Frank, Wilson, & Brown, 2001; Brown, Frank, Tang, Quirk, & Wilson, 1998; Kass & Ventura, 2001; Metzner, Koch, Wessel, & Gabbiani, 1998; Zhang, Ginzburg, McNaughton, & Sejnowski, 1998). We propose here to estimate the entropy rate of a spike train from an inhomogeneous hidden Markov model of the spike intervals. The model is constructed by building a context tree structure to lay out the conditional probabilities of various subsequences of the spike train. For each state in the Markov chain, we assume a gamma distribution over the spike intervals, although any appropriate distribution may be employed as circumstances dictate. The entropy and confidence intervals for the entropy are calculated from bootstrapping samples taken from a large raw data sequence. The estimator was first tested on synthetic data generated by multiple-order Markov chains, and it always converged to the theoretical Shannon entropy rate (except in the case of a sixth-order model, where the calculations were terminated before convergence was reached). We also applied the method to experimental data and compare its performance with that of several other methods of entropy estimation.

We report on a search for the standard model Higgs boson produced in association with a W or Z boson in p (p) over bar collisions at root s = 1.96 TeV recorded by the CDF II experiment at the Tevatron in a data sample corresponding to an integrated luminosity of 2.1 fb(-1). We consider events which have no identified charged leptons, an imbalance in transverse momentum, and two or three jets where at least one jet is consistent with originating from the decay of a b hadron. We find good agreement between data and background predictions. We place 95% confidence level upper limits on the production cross section for several Higgs boson masses ranging from 110 GeV/c(2) to 150 GeV/c(2). For a mass of 115 GeV/c(2) the observed (expected) limit is 6.9 (5.6) times the standard model prediction.

Background: Our objective was to develop a consistent molecular definition of psoriasis. There have been several published microarray studies of psoriasis, and we compared disease-related genes identified across these different studies of psoriasis with our own in order to establish a consensus. Methodology/Principal Findings: We present a psoriasis transcriptome from a group of 15 patients enrolled in a clinical study, and assessed its biological validity using a set of important pathways known to be involved in psoriasis. We also identified a key set of cytokines that are now strongly implicated in driving disease-related pathology, but which are not detected well on gene array platforms and require more sensitive methods to measure mRNA levels in skin tissues. Comparison of our transcriptome with three other published lists of psoriasis genes showed apparent inconsistencies based on the number of overlapping genes. We extended the well-established approach of Gene Set Enrichment Analysis (GSEA) to compare a new study with these other published list of differentially expressed genes (DEG) in a more comprehensive manner. We applied our method to these three published psoriasis transcriptomes and found them to be in good agreement with our study. Conclusions/Significance: Due to wide variability in clinical protocols, platform and sample handling, and subtle disease-related signals, intersection of published DEG lists was unable to establish consensus between studies. In order to leverage the power of multiple transcriptomes reported by several laboratories using different patients and protocols, more sophisticated methods like the extension of GSEA presented here, should be used in order to overcome the shortcomings of overlapping individual DEG approach.

Voltage sensors regulate the conformations of voltage-dependent ion channels and enzymes. Their nearly switchlike response as a function of membrane voltage comes from the movement of positively charged amino acids, arginine or lysine, across the membrane field. We used mutations with natural and unnatural amino acids, electrophysiological recordings, and x-ray crystallography to identify a charge transfer center in voltage sensors that facilitates this movement. This center consists of a rigid cyclic "cap" and two negatively charged amino acids to interact with a positive charge. Specific mutations induce a preference for lysine relative to arginine. By placing lysine at specific locations, the voltage sensor can be stabilized in different conformations, which enables a dissection of voltage sensor movements and their relation to ion channel opening.