DISC1 at 10: connecting psychiatric genetics and neuroscience

Psychiatric genetics research, as exemplified by the DISC1 gene, aspires to inform on mental health etiology and to suggest improved strategies for intervention. DISC1 was discovered in 2000 through the molecular cloning of a chromosomal translocation that segregated with a spectrum of major mental illnesses in a single large Scottish family. Through in vitro experiments and mouse models, DISC1 has been firmly established as a genetic risk factor for a spectrum of psychiatric illness. As a consequence of its protein scaffold function, the DISC1 protein impacts on many aspects of brain function, including neurosignaling and neurodevelopment. DISC1 is a pathfinder for understanding psychopathology, brain development, signaling and circuitry. Although much remains to be learnt and understood, potential targets for drug development are starting to emerge, and in this review, we will discuss the 10 years of research that has helped us understand key roles of DISC1 in psychiatric disease.     

Shotgun proteomic analysis of the unicellular alga Ostreococcus tauri

Ostreococcus tauri is a unicellular green alga and amongst the smallest and simplest free-living eukaryotes. The O. tauri genome sequence was determined in 2006. Molecular, physiological and taxonomic data that has been generated since then highlight its potential as a simple model species for algae and plants. However, its proteome remains largely unexplored. This paper describes the global proteomic study of O. tauri, using mass spectrometry-based approaches: phosphopeptide enrichment, cellular fractionation, label-free quantification and (15)N metabolic labeling. The O. tauri proteome was analyzed under the following conditions: sampling at different times during the circadian cycle, after 24h of illumination, after 24h of darkness and under various nitrogen source supply levels. Cell cycle related proteins such as dynamin and kinesin were significantly up-regulated during the daylight-to-darkness transition. This is reflected by their higher intensity at ZT13 and this transition phase coincides with the end of mitosis. Proteins involved in several metabolic mechanisms were found to be up-regulated under low nitrogen conditions, including carbon storage pathways, glycolysis, phosphate transport, and the synthesis of inorganic polyphosphates. Ostreococcus tauri responds to low nitrogen conditions by reducing its nitrogen assimilation machinery which suggests an atypical adaptation mechanism for coping with a nutrient-limited environment.     

Combining experimental and predicted datasets for determination of the subcellular location of proteins in Arabidopsis

Substantial experimental datasets defining the subcellular location of Arabidopsis (Arabidopsis thaliana) proteins have been reported in the literature in the form of organelle proteomes built from mass spectrometry data (approximately 2,500 proteins). Subcellular location for specific proteins has also been published based on imaging of chimeric fluorescent fusion proteins in intact cells (approximately 900 proteins). Further, the more diverse history of biochemical determination of subcellular location is stored in the entries of the Swiss-Prot database for the products of many Arabidopsis genes (approximately 1,800 proteins). Combined with the range of bioinformatic targeting prediction tools and comparative genomic analysis, these experimental datasets provide a powerful basis for defining the final location of proteins within the wide variety of subcellular structures present inside Arabidopsis cells. We have analyzed these published experimental and prediction data to answer a range of substantial questions facing researchers about the veracity of these approaches to determining protein location and their interrelatedness. We have merged these data to form the subcellular location database for Arabidopsis proteins (SUBA), providing an integrated understanding of protein location, encompassing the plastid, mitochondrion, peroxisome, nucleus, plasma membrane, endoplasmic reticulum, vacuole, Golgi, cytoskeleton structures, and cytosol (www.suba.bcs.uwa.edu.au). This includes data on more than 4,400 nonredundant Arabidopsis protein sequences. We also provide researchers with an online resource that may be used to query protein sets or protein families and determine whether predicted or experimental location data exist; to analyze the nature of contamination between published proteome sets; and/or for building theoretical subcellular proteomes in Arabidopsis using the latest experimental data.     

Wnt/beta-catenin signaling acts upstream of N-myc, BMP4, and FGF signaling to regulate proximal-distal patterning in the lung

Branching morphogenesis in the lung serves as a model for the complex patterning that is reiterated in multiple organs throughout development. Beta-catenin and Wnt signaling mediate critical functions in cell fate specification and differentiation, but specific functions during branching morphogenesis have remained unclear. Here, we show that Wnt/beta-catenin signaling regulates proximal-distal differentiation of airway epithelium. Inhibition of Wnt/beta-catenin signaling, either by expression of Dkk1 or by tissue-specific deletion of beta-catenin, results in disruption of distal airway development and expansion of proximal airways. Wnt/beta-catenin functions upstream of BMP4, FGF signaling, and N-myc. Moreover, we show that beta-catenin and LEF/TCF activate the promoters of BMP4 and N-myc. Thus, Wnt/beta-catenin signaling is a critical upstream regulator of proximal-distal patterning in the lung, in part, through regulation of N-myc, BMP4, and FGF signaling.     

Mutations remote from the human gonadotropin-releasing hormone (GnRH) receptor-binding sites specifically increase binding affinity for GnRH II but not GnRH I: evidence for ligand-selective, receptor-active conformations

The human gonadotropin-releasing hormone (GnRH) receptor is evolutionarily configured for high affinity binding of GnRH I ([Tyr(5),Leu(7),Arg(8)]GnRH) but at lower affinity for GnRH II ([His(5),Trp(7),Tyr(8)]GnRH). GnRH I is more potent in the activation of the G(q/11) protein in the gonadotrope; however, GnRH II is more potent in the stimulation of apoptosis and antiproliferative effects through activating G(i) protein-mediated signaling, implying that GnRH I and II selectively stabilize different receptor-active conformations that preferentially couple to different signaling pathways. Receptor activation involves ligand induction or conformational selection, but the molecular basis of the communication between ligand-binding sites and receptor allosteric sites remains unclear. We have sought conformational coupling between receptor-ligand intermolecular interactions and intramolecular interaction networks in the human GnRH receptor by mutating remote residues that induce differential ligand binding affinity shifts for GnRH I and II. We have demonstrated that certain Ala mutations in the intracellular segments of transmembrane domains 3 (Met(132)), 5 (Met(227)), 6 (Phe(272) and Phe(276)), and 7 (Ile(322) and Tyr(323)) of the human GnRH receptor allosterically increased ligand binding affinity for GnRH II but had little effect on GnRH I binding affinity. We examined the role of the three amino acids that differ in these two ligands, and we found that Tyr(8) in GnRH II plays a dominant role for the increased affinity of the receptor mutants for GnRH II. We propose that creation of a high affinity binding site for GnRH II accompanies receptor conformational changes, i.e."induced fit" or "conformational selection," mainly determined by the intermolecular interactions between Tyr(8) and the receptor contact residues, which can be facilitated by disruption of particular sets of receptor-stabilizing intramolecular interactions. The findings suggest that GnRH I and II binding may selectively stabilize different receptor-active conformations and therefore different ligand-induced selective signaling described previously for these ligands.     

Is a large-scale DNA-based inventory of ancient life possible?

A complete DNA-based inventory of the Earth's present biota using large-scale high-throughput DNA sequencing of signature region(s) (DNA barcoding) is an ambitious proposal rivaling the Human Genome Project. We examine whether this approach will also enable us to assess the past diversity of the earth's biota. To test this, we sequenced the 5' terminus of the mitochondrial cytochrome c oxidase I (COI) gene of individuals belonging to a group of extinct ratite birds, the moa of New Zealand. Moa comprised a large number of taxa that radiated in isolation on this oceanic landmass. Using a phylogenetic approach based on a large data set including protein coding and 12S DNA sequences as well as morphology, we now have precise information about the number of moa species that once existed. We show that each of the moa species detected using this extensive data set has a unique COI barcode(s) and that they all show low levels of within-species COI variation. Consequently, we conclude that COI sequences accurately identify the species discovered using the larger data set. Hence, more generally, this study suggests that DNA barcoding might also help us detect other extinct animal species and that a large-scale inventory of ancient life is possible.     

On the solution structure of the T4 sliding clamp (gp45)

Examination by time-resolved fluorescence spectroscopy of the trimeric bacteriophage T4 clamp protein labeled across its three subunit interfaces with a fluorescence resonance energy transfer (FRET) pair indicates that the clamp exists in just one state in solution, with one open and two closed interfaces. This is in contrast to what is observed in the X-ray crystal structure. The population distribution of the trFRET distance is bimodal, giving 67% as 17 A and 33% as 42 A. This leads to the conclusion that gp45 exists in an asymmetric open state in solution. The further increase in the separation of the FRET pair in the presence of the clamp loader and ATP may be ascribed to either further opening of the open interface or the opening of a closed interface. The ramifications for replisome remodeling by this pathway are discussed.     

Sensitivity of Bayes estimators to hyper-parameters with an application to maximum yield from fisheries

Priors are seldom unequivocal and an important component of Bayesian modeling is assessment of the sensitivity of the posterior to the specified prior distribution. This is especially true in fisheries science where the Bayesian approach has been promoted as a rigorous method for including existing information from previous surveys and from related stocks or species. These informative priors may be highly contested by various interest groups. Here, formulae for the first and second derivatives of Bayes estimators with respect to hyper-parameters of the joint prior density are given. The formula for the second derivative provides a correction to a previously published result. The formulae are shown to reduce to very convenient and easily implemented forms when the hyper-parameters are for exponential family marginal priors. For model parameters with such priors it is shown that the ratio of posterior variance to prior variance can be interpreted as the sensitivity of the posterior mean to the prior mean. This methodology is applied to a nonlinear state-space model for the biomass of South Atlantic albacore tuna and sensitivity of the maximum sustainable yield to the prior specification is examined.     

The wild-type circadian period of Neurospora is encoded in the residual network of the null frq mutants

We model theoretically the response of the widely studied circadian oscillator of Neurospora crassa to inactivation of the frq gene. The resulting organism has been termed "arrhythmic" under constant conditions. Under entrainment to periodic temperature cycles Roenneberg, Merrow and coworkers have shown that the phase angle at which spore formation occurs depends on the entrainment period, curiously even in the null frq mutants (frq9 and frq10). We show that such a response does not imply the presence of a self-sustained free-running oscillator. We derive a simple candidate model (a damped harmonic oscillator) for the null frq mutants that successfully reproduces the observed phase angle response. An endogenous period of 21 h for the damped harmonic oscillator coincides with the endogenous period of wild-type Neurospora. This suggests that the (noise driven) "residual system" present in the mutants may have a significant timekeeping role in the wild-type organism. Our model (with no change of parameters) was then used to investigate spore formation patterns under constant conditions and reproduces the corresponding experimental data of Aronson et al. (Proc. Natl. Acad. Sci. USA 91 (1994) 7683.)