Systems biology of mammalian circadian clocks

Systems biology of mammalian circadian clocks

S22 Abstracts Post-traumatic stress disorder (PTSD) is triggered by a single overwhelmingly traumatic, often life-threatening, event. Some component...

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S22

Abstracts

Post-traumatic stress disorder (PTSD) is triggered by a single overwhelmingly traumatic, often life-threatening, event. Some components of the fear response in PTSD persist well beyond the original event. However, current animal models of PTSD have focused on the immediate effects of prolonged stress on cognitive function, without providing insights into the cellular mechanisms underlying the emotional symptoms of PTSD. The present study captures some of the defining features of PTSD in human patients using an animal model wherein a single episode of acute stress (2 hours) causes higher anxiety and spinogenesis in the amygdala not one, but ten days later. Whole-cell recordings in rat brain slices of the lateral amygdala (LA) show that stress lowers the frequency of miniature GABAergic inhibitory currents (mIPSCs) only 10 days, but not 1 day, after stress. Taken together, our findings provide an attractive new animal model for studying synaptic mechanisms in the amygdala that may underlie key affective symptoms of PTSD. doi:10.1016/j.neures.2009.09.1608

SY2-E3-4 Phenotypic abnormalities caused by genetic loss of PSD95 (Dlg4) reveal a potential role for MAGUKs in autism and Williams Syndrome Andrew Holmes NIAAA, USA Genes regulating synaptic function and learning are implicated in neurodevelopmental disorders. The membrane-associated guanylate kinases (MAGUK), including PSD-95 (DLG4), orchestrate protein-protein interactions and receptor stabilization at glutamatergic synapses, and are key mediators of synaptic plasticity. Genetic variation in the MAGUKs DLG1 and DLG3 are associated with autistic-like traits and X-linked mental retardation. To examine a possible role for DLG4 in neurodevelopmental disorders, we phenotypically characterized Dlg4 deletion in the mouse. Dlg4 knockouts phenocopy the principle behavioral symptoms of the neurodevelopmental disorder Williams syndrome (WS), including heightened anxiety-like behavior, abnormal social behavior and motor incoordination. Genome-wide analysis of gene expression revealed alterations in the forebrain expression of a network of key synaptic proteins, as well as a 50% downregulation of the WS candidate gene, Cyln2. These findings reveal a potential role for DLG4 in WS and, more generally, provide support for the characterization of neurodevelopmental disorders as synaptopathies. doi:10.1016/j.neures.2009.09.1609

SY2-E3-5 Immature dentate gyrus as a candidate endophenotype of psychiatric disorders Tsuyoshi Miyakawa

factors. We propose that differentiating ES cells are a powerful tool for investigating the fundamental mechanisms of the robust mammalian circadian clock system. doi:10.1016/j.neures.2009.09.1611

SY2-F1-2 Phosphorylation-dependent regulation of CLOCK-BMAL1 function in circadian clockwork Hikari Yoshitane, Yoshitaka Fukada Department Biophys. and Biochem., Grad. Sch. Sci., University Tokyo, Japan In mammalian circadian clockwork, CLOCK-BMAL1 heterodimer activates E-boxdependent transcription, while its activity is suppressed by circadian binding with negative regulators. Here we found that CLOCK is hyperphosphorylated in the suppression phase of E-box-dependent transcription in the mouse liver and NIH3T3 cells. In vivo phosphorylation sites of CLOCK were identified by MS/MS analysis and Asp-mutations at the sites markedly weakened the transactivation ability of CLOCK-BMAL1 by decreasing the nuclear amount of CLOCK protein and inhibiting the DNA-binding activity. Coexpression of a negative regulator CIPC in NIH3T3 cells significantly promoted CLOCK phosphorylation, while deletion mutant CLOCK lacking CIPC-binding domain was far less phosphorylated and much more stabilized than wild-type CLOCK in vivo. On the other hand, promotion of CLOCK phosphorylation facilitated its proteasomal degradation. Collectively, CLOCK phosphorylation contributes to suppression of CLOCK-BMAL1-mediated transactivation through dual regulation; inhibition of CLOCK activity and promotion of its degradation. doi:10.1016/j.neures.2009.09.1612

SY2-F1-3 KaiC defines circadian clock of cyanobacteria Takao Kondo Nagoya University, Japan We studied the KaiC phosphorylation cycle in vitro and confirmed that the ATPase activity of KaiC defines the period length and its temperature compensation. KaiC possesses extremely weak but stable ATPase activity (15 molecules of ATP per day), and the addition of KaiA and KaiB makes the activity oscillate with a circadian period in vitro. The ATPase activity of KaiC is inherently temperature-invariant, suggesting that temperature compensation of the circadian period could be driven by simple ATPase reaction. Interestingly, the activities of wild-type KaiC and five period-mutant proteins are directly proportional to their in vivo circadian frequencies, indicating that the ATPase activity defines the circadian period. We propose that KaiC ATPase activity constitutes the most fundamental reaction underlying circadian periodicity in cyanobacteria. doi:10.1016/j.neures.2009.09.1613

ICMS, Fujita Health University, Toyoake, Japan Elucidating the neural and genetic factors underlying psychiatric illness is hampered by current methods of clinical diagnosis. The identification and investigation of clinical endophenotypes may be one solution, but represents a considerable challenge in human subjects. Previously, we reported that mice heterozygous for a null mutation of a-CaMKII (a-CaMKII+/-), a key molecule in synaptic plasticity, have profoundly dysregulated behaviors including a severe working memory deficit, which is an endophenotype of schizophrenia and other psychiatric disorders. In addition, we found that almost all the neurons in the dentate gyrus (DG) of the mutant mice failed to mature at molecular, morphological and electrophysiological levels. Here I show that the mice lacking a transcription factor,Schnurri, exhibit abnormal behavioral pattern that is similar to that of a-CaMKII+/- mice. Schnurri KO mice Potential mechanisms underlying the “immature dentate gyrus” and strategies to normalize this phenotype will be also discussed. doi:10.1016/j.neures.2009.09.1610

SY2-F1-1 Chronogenesis during the ES cell differentiation in vitro Kazuhiro Yagita

SY2-F1-4 Systems biology of mammalian circadian clocks Hiroki Ueda RIKEN, CDB, Japan Mammalian circadian clock system is a complex and dynamic system consisting of complicatedly integrated regulatory loops and displaying the various dynamic behaviors including (i) endogenous oscillation with about 24-h period, (ii) entrainment to the external environmental changes (temperature and light cycle), and (iii) temperature compensation over the wide range of temperature. The logic of such biological networks such as circadian clocks is difficult to elucidate without (1) comprehensive identification of network structure (Ueda et al., 2002, 2005; Sato et al., 2006), (2) prediction and validation based on quantitative measurement and perturbation of network behavior (Ukai et al., 2007), and (3) design and implementation of artificial networks of identified structure and observed dynamics (Ukai-Tadenuma et al., 2008). In this symposium, we will report on the current progress in the analysis and synthesis of mammalian circadian clocks. References

Osaka University Graduate School of Medicine, Japan The molecular oscillations underlying the generation of circadian rhythmicity in mammals develop gradually during ontogenesis. The process of chronogenesis in embryonic cells includes essential events that generate the self-sustaining and robust circadian oscillator. Here, we show that the mammalian circadian oscillator develops during the differentiation of mouse embryonic stem (ES) cells without maternal or external entraining factors. In ES cells, no circadian bioluminescence oscillation was observed. On the other hands, circadian cycles of bioluminescence gradually developed when the cells were treated with all-trans-retinoic acid (RA) to induce differentiation. These results suggest that the circadian oscillator in mammals is acquired during differentiation without the assist of maternal entrainment

Sato, T.K., et al., 2006. Nat. Genet. 38, 312–319. Ueda, H.R., et al., 2002. Nature 418, 534–539. Ueda, H.R., et al., 2005. Nat. Genet. 37, 187–192. Ukai, H., et al., 2007. Nat. Cell Biol. 9, 1327–1334. Ukai-Tadenuma, M., et al., 2008. Nat. Cell Biol. 10, 1154–1163. doi:10.1016/j.neures.2009.09.1614