Kun Ho Lee

Adrenal peripheral clock controls the autonomous circadian rhythm of glucocorticoid by causing rhythmic steroid production

Glucocorticoid (GC) is an adrenal steroid with diverse physiological effects. It undergoes a robust daily oscillation, which has been thought to be driven by the master circadian clock in the suprachiasmatic nucleus of the hypothalamus via the hypothalamus–pituitary–adrenal axis. However, we show that the adrenal gland has its own clock and that the peripheral clockwork is tightly linked to steroidogenesis by the steroidogenic acute regulatory protein. Examination of mice with adrenal-specific knockdown of the canonical clock protein BMAL1 reveals that the adrenal clock machinery is required for circadian GC production. Furthermore, behavioral rhythmicity is drastically affected in these animals, together with altered expression of Period1, but not Period2, in several peripheral organs. We conclude that the adrenal peripheral clock plays an essential role in harmonizing the mammalian circadian timing system by generating a robust circadian GC rhythm.

Neural correlates of superior intelligence: Stronger recruitment of posterior parietal cortex

General intelligence (g) is a common factor in diverse cognitive abilities and a major influence on life outcomes. Neuroimaging studies in adults suggest that the lateral prefrontal and parietal cortices play a crucial role in related cognitive activities including fluid reasoning, the control of attention, and working memory. Here, we investigated the neural bases for intellectual giftedness (superior-g) in adolescents, using fMRI. The participants consisted of a superior-g group (n = 18, mean RAPM = 33.9 +/- 0.8, >99%) from the national academy for gifted adolescents and the control group (n = 18, mean RAPM = 22.8 +/- 1.6, 60%) from local high schools in Korea (mean age = 16.5 +/- 0.8). fMRI data were acquired while they performed two reasoning tasks with high and low g-loadings. In both groups, the high g-loaded tasks specifically increased regional activity in the bilateral fronto-parietal network including the lateral prefrontal, anterior cingulate, and posterior parietal cortices. However, the regional activations of the superior-g group were significantly stronger than those of the control group, especially in the posterior parietal cortex. Moreover, regression analysis revealed that activity of the superior and intraparietal cortices (BA 7/40) strongly covaried with individual differences in g (r = 0.71 to 0.81). A correlated vectors analysis implicated bilateral posterior parietal areas in g. These results suggest that superior-g may not be due to the recruitment of additional brain regions but to the functional facilitation of the fronto-parietal network particularly driven by the posterior parietal activation.

Maternal Stress Produces Learning Deficits Associated with Impairment of NMDA Receptor-Mediated Synaptic Plasticity

Stress in adulthood can have a profound effect on physiology and behavior, but the extent to which prolonged maternal stress affects the brain function of offspring when they are adult remains primarily unknown. In the present work, chronic immobilization stress to pregnant mice affected fetal growth and development. When pups born from stressed mice were reared to adulthood in an environment identical to that of nonstressed controls, several physiological parameters were essentially unaltered. However, spatial learning and memory was significantly impaired in the maternally stressed offspring in adulthood. Furthermore, electrophysiological examination revealed a significant reduction in NMDA receptor-mediated long-term potentiation in the CA1 area of hippocampal slices. Subsequent biochemical analysis demonstrated a substantial decrease in NR1 and NR2B subunits of the NMDA receptor in synapses of the hippocampus, and the interaction between these two subunits appeared to be reduced. These results suggest that prolonged maternal stress leads to long-lasting malfunction of the hippocampus, which extends to and is manifested in adulthood.

Multiple Bases of Human Intelligence Revealed by Cortical Thickness and Neural Activation

We hypothesized that individual differences in intelligence (Spearmans g) are supported by multiple brain regions, and in particular that fluid (gF) and crystallized (gC) components of intelligence are related to brain function and structure with a distinct profile of association across brain regions. In 225 healthy young adults scanned with structural and functional magnetic resonance imaging sequences, regions of interest (ROIs) were defined on the basis of a correlation between g and either brain structure or brain function. In these ROIs, gC was more strongly related to structure (cortical thickness) than function, whereas gF was more strongly related to function (blood oxygenation level-dependent signal during reasoning) than structure. We further validated this finding by generating a neurometric prediction model of intelligence quotient (IQ) that explained 50% of variance in IQ in an independent sample. The data compel a nuanced view of the neurobiology of intelligence, providing the most persuasive evidence to date for theories emphasizing multiple distributed brain regions differing in function.

Dual Modification of BMAL1 by SUMO2/3 and Ubiquitin Promotes Circadian Activation of the CLOCK/BMAL1 Complex

ABSTRACT Heterodimers of BMAL1 and CLOCK drive rhythmic expression of clock-controlled genes, thereby generating circadian physiology and behavior. Posttranslational modifications of BMAL1 play a key role in modulating the transcriptional activity of the CLOCK/BMAL1 complex during the circadian cycle. Recently, we demonstrated that circadian activation of the heterodimeric transcription factor is accompanied by ubiquitin-dependent proteolysis of BMAL1. Here we show that modification by SUMO localizes BMAL1 exclusively to the promyelocytic leukemia nuclear body (NB) and simultaneously promotes its transactivation and ubiquitin-dependent degradation. Under physiological conditions, BMAL1 was predominantly conjugated to poly-SUMO2/3 rather than SUMO1, and the level of these conjugates underwent rhythmic variation, peaking at times of maximum E-box-mediated circadian transcription. Interestingly, mutation of the sumoylation site (Lys259) of BMAL1 markedly inhibited both its ubiquitination and its proteasome-mediated proteolysis, and these effects were reversed by covalent attachment of SUMO3 to the C terminus of the mutant BMAL1. Consistent with this, SUSP1, a SUMO protease highly specific for SUMO2/3, abolished ubiquitination, as well as sumoylation of BMAL1, while the ubiquitin protease UBP41 blocked BMAL1 ubiquitination but induced accumulation of polysumoylated BMAL1 and its localization to the NB. Furthermore, inhibition of proteasome with MG132 elicited robust nuclear accumulation of SUMO2/3- and ubiquitin-modified BMAL1 that was restricted to the transcriptionally active stage of the circadian cycle. These results indicate that dual modification of BMAL1 by SUMO2/3 and ubiquitin is essential for circadian activation and degradation of the CLOCK/BMAL1 complex.

Mitochondrial membrane depolarization and the selective death of dopaminergic neurons by rotenone: protective effect of coenzyme Q10

Chronic exposure to the pesticide rotenone induces a selective degeneration of nigrostriatal dopaminergic neurons and reproduces the features of Parkinsons disease in experimental animals. This action is thought to be relevant to its inhibition of the mitochondrial complex I, but the precise mechanism of this suppression in selective neuronal death is still elusive. Here we investigate the mechanism of dopaminergic neuronal death mediated by rotenone in primary rat mesencephalic neurons. Low concentrations of rotenone (5–10 nm) induce the selective death of dopaminergic neurons without significant toxic effects on other mesencephalic cells. This cell death was coincident with apoptotic events including capsase‐3 activation, DNA fragmentation, and mitochondrial membrane depolarization. Pretreatment with coenzyme Q10, the electron transporter in the mitochondrial respiratory chain, remarkably reduced apoptosis as well as the mitochondrial depolarization induced by rotenone, but other free radical scavengers such as N‐acetylcysteine, glutathione, and vitamin C did not. Furthermore, the selective neurotoxicity of rotenone was mimicked by the mitochondrial protonophore carbonyl cyanide 4‐(trifluoromethoxy) phenylhydrazone (FCCP), a cyanide analog that effectively collapses a mitochondrial membrane potential. These data suggest that mitochondrial depolarization may play a crucial role in rotenone‐induced selective apoptosis in rat primary dopaminergic neurons.

BMAL1 Shuttling Controls Transactivation and Degradation of the CLOCK/BMAL1 Heterodimer

ABSTRACT CLOCK and BMAL1 are bHLH-PAS-containing transcription factors that bind to E-box elements and are indispensable for expression of core circadian clock components such as the Per and Cry genes. A key step in expression is the heterodimerization of CLOCK and BMAL1 and their accumulation in the nucleus with an approximately 24-h periodicity. We show here that nucleocytoplasmic shuttling of BMAL1 is essential for transactivation and for degradation of the CLOCK/BMAL1 heterodimer. Using serial deletions and point mutants, we identified a functional nuclear localization signal and Crm1-dependent nuclear export signals in BMAL1. Transient-transfection experiments revealed that heterodimerization of CLOCK and BMAL1 accelerates their turnover, as well as E-box-dependent clock gene transcription. Moreover, in embryonic mouse fibroblasts, robust transcription of Per2 is tightly associated with massive degradation of the CLOCK/BMAL1 heterodimer. CRY proteins suppressed this process during the transcription-negative phase and led to nuclear accumulation of the CLOCK/BMAL1 heterodimer. Thus, these findings suggest that the decrease of BMAL1 abundance during the circadian cycle reflects robust transcriptional activation of clock genes rather than inhibition of BMAL1 synthesis.

Rapid activation of CLOCK by Ca2+‐dependent protein kinase C mediates resetting of the mammalian circadian clock

In mammals, immediate‐early transcription of the Period 1 (Per1) gene is crucial for resetting the mammalian circadian clock. Here, we show that CLOCK is a real signalling molecule that mediates the serum‐evoked rapid induction of Per1 in fibroblasts through the Ca2+‐dependent protein kinase C (PKC) pathway. Stimulation with serum rapidly induced nuclear translocation, heterodimerization and Ser/Thr phosphorylation of CLOCK just before the surge of Per1 transcription. Serum‐induced CLOCK phosphorylation was abolished by treatment with PKC inhibitors but not by other kinase inhibitors. Consistently, the interaction between CLOCK and PKC was markedly increased shortly after serum shock, and the Ca2+‐dependent PKC isoforms PKCα and PKCγ phosphorylated CLOCK in vitro. Furthermore, phorbol myristic acetate treatment triggered immediate‐early transcription of Per1 and also CLOCK phosphorylation, which were blocked by a Ca2+‐dependent PKC inhibitor. These findings indicate that CLOCK activation through the Ca2+‐dependent PKC pathway might have a substantial role in phase resetting of the circadian clock.

Neuregulin Stimulates Myogenic Differentiation in an Autocrine Manner

During myogenesis, mononucleated myoblasts form multinucleated myotubes by membrane fusion. Efficiency of this intercellular process can be maximized by a simultaneous progress, with a time window, of other neighboring myoblasts in the differentiation program. This phenomenon has been described as the community effect. It proposes the existence of a molecule that acts as a differentiation-inducing signal to a group of identical cells. Here ,we show that neuregulin is a strong candidate for this molecule in myoblast differentiation. The expression of neuregulin increased rapidly but transiently at early stage of differentiation of rat L6 cells. Neuregulin showed a potent differentiation-promoting activity in membrane fusion and expression of myosin heavy chain. The antibodies raised against neuregulin and its cognate receptor ErbB3, which were capable of neutralizing the signal pathway, inhibited myotube formation and expression of myosin heavy chain in both L6 cells and primary rat myoblasts. The progress of differentiation was mostly halted after the expression of myogenin and cell cycle arrest. These results suggest that the activation of an autocrine signaling of neuregulin may provide a basic mechanism for the community effect observed in the differentiation of the embryonic muscle cells.

Coactivation of the CLOCK–BMAL1 complex by CBP mediates resetting of the circadian clock

The transcription factor CLOCK–BMAL1 is a core component of the molecular clock machinery that drives circadian gene expression and physiology in mammals. Recently, we reported that this heterodimeric transcription factor functions as a signaling molecule in response to the resetting stimuli via the Ca2+-dependent protein kinase C pathway. Here, we demonstrate that the CREB-binding protein (CBP) plays a key role in rapid activation of the CLOCK–BMAL1 heterodimer that leads to phase resetting of the circadian clock. Under physiological conditions, a bimolecular fluorescence complementation (BiFC) assay revealed that CLOCK and BMAL1 dimerize in the cytoplasm and subsequently translocate into the nucleus in response to serum stimuli (mean time duration was 29.2 minutes and mean velocity 0.7 μm/minute). Concomitantly, BMAL1 rapidly recruited CBP on Per1 promoter E-box, but not p300 (a functional analog of CBP), in the discrete nuclear foci. However, recruitment of CBP by cAMP/Ca2+ response element-binding (CREB) protein on CRE was not markedly increased upon delivery of the resetting stimuli. Furthermore, overexpression of CBP greatly potentiated the CLOCK–BMAL1-mediated Per1 transcription, and this effect was completely abolished by site-directed mutation of E-box elements, but not by the mutation of CRE in the Per1 promoter. Furthermore, molecular knockdown of CBP severely dampened circadian oscillation of clock gene expression triggered by the resetting stimuli. These findings suggest that CBP recruitment by BMAL1 mediates acute transactivation of CLOCK–BMAL1, thereby inducing immediate-early Per1 transcription and phase resetting of the circadian clock.