Soontaek Hong

Expression of thymosin β in the rat brain following transient global ischemia

Thymosin beta (Tbeta) isoforms play an important role in the organization of the cytoskeleton by sequestering G-actin during development of the mammalian brain. In this study, we examined changes in the expression of Tbeta4 and Tbeta15 after transient global ischemia. Tbeta15 mRNA increased gradually in the dentate gyrus (DG) of the hippocampal formation from 3 h after reperfusion and peaked 9 h later. Similarly, a significant increase in Tbeta4 mRNA level was observed in the DG 12 h after reperfusion. Tbeta4 and Tbeta15 proteins were found in different cell types in control brains; Tbeta15 was expressed in a subset of doublecortin (DCX)-positive cells in the DG, whereas Tbeta4-IR was observed in DG neurons and nearby microglial cells. After ischemia, Tbeta15-IR was found in DG neurons and Tbeta4-IR in the reactivated microglial cells. Interestingly, Tbeta15-IR accumulated in the nuclei of CA1 neurons, which are vulnerable to ischemic insults. These results suggest that Tbeta4 and Tbeta15 function in different cellular contexts during ischemia-induced responses.

Preso regulation of dendritic outgrowth through PI(4,5)P2-dependent PDZ interaction with βPix

In neuronal development, dendritic outgrowth and arborization are important for the establishment of neural circuit formation. A previous study reported that PSD‐95‐interacting regulator of spine morphogenesis (Preso) formed a complex with PAK‐interacting exchange factor‐beta (βPix) via PSD‐95/Dlg/ZO‐1 (PDZ) interaction. Here, we report that Preso and its binding protein, βPix, are localized in dendritic growth cones. Knockdown and dominant‐negative inhibition of Preso in cultured neurons markedly reduced the dendritic outgrowth but not branching, and led to a decrease in the intensity of βPix and F‐actin in neuronal dendritic tips. Moreover, phosphatidylinositol 4,5‐bisphosphate (PIP2) induced a conformational change in Preso toward the open PDZ domain and enhanced the interaction with βPix. In addition, the Preso band 4.1 protein, ezrin, radixin and moesin (FERM) domain mutant is unable to interact with PIP2 and it did not rescue the Preso‐knockdown effect. These results indicate that PIP2 is a key signalling molecule that regulates dendritic outgrowth through activation of small GTPase signalling via interaction between Preso and βPix.

Hes6 Controls Cell Proliferation via Interaction with cAMP-response Element-binding Protein-binding Protein in the Promyelocytic Leukemia Nuclear Body

Hes6 is a basic helix-loop-helix transcription factor that functions in the differentiation of pluripotent progenitor cells and during tumorigenesis. However, the molecular mechanism for its function is largely unknown. Here we show that Hes6 is a component of the promyelocytic leukemia nuclear body (PML-NB) complex in the nuclei and that Hes6 inhibits cell proliferation through induction of p21 cyclin-dependent kinase inhibitor. We further show that Hes6 directly interacts with CREB-binding protein (CBP), one of the key components of PML-NB, via its basic domain. This association is critical for p21 induction through multiple mechanisms, including chromatin remodeling and p53 acetylation. Taken together, these results suggest that the Hes6-CBP complex in PML-NB may influence the proliferation of cells via p53-dependent and -independent pathways.

Altered expression of synaptotagmin 13 mRNA in adult mouse brain after contextual fear conditioning

Contextual fear memory processing requires coordinated changes in neuronal activity and molecular networks within brain. A large number of fear memory-related genes, however, still remain to be identified. Synaptotagmin 13 (Syt13), an atypical member of synaptotagmin family, is highly expressed in brain, but its functional roles within brain have not yet been clarified. Here, we report that the expression of Syt13 mRNA in adult mouse brain was altered following contextual fear conditioning. C57BL/6 mice were exposed to a novel context and stimulated by strong electrical footshock according to a contextual fear conditioning protocol. After 24 h, the mice were re-exposed to the context without electrical footshock for the retrieval of contextual fear memory. To investigate the relationship between Syt13 and contextual fear memory, we carried out in situ hybridization and analyzed gene expression patterns for Syt13 at four groups representing temporal changes in brain activity during contextual fear memory formation. Contextual fear conditioning test induced significant changes in mRNA levels for Syt13 within various brain regions, including lateral amygdala, somatosensory cortex, piriform cortex, habenula, thalamus, and hypothalamus, during both acquisition and retrieval sessions. Our data suggest that Syt13 may be involved in the process of contextual fear memory.

Inositol 1,4,5 trisphosphate 3-kinase A is a novel microtubule-associated protein: PKA-dependent phosphoregulation of microtubule binding affinity

Background: IP3K-A mediates synaptic plasticity and memory formation. However, its molecular mechanism is not clear. Results: Tubulin is identified as an IP3K-A-binding protein, and PKA reduces the microtubule binding affinity of IP3K-A. Conclusion: IP3K-A is a novel microtubule-associated protein. PKA-dependent phosphorylation negatively regulates interactions between IP3K-A and microtubule. Significance: Phosphoregulation of IP3K-A may serve as its novel regulatory mechanism. Inositol 1,4,5-trisphosphate 3-kinase A (IP3K-A) is a brain specific and F-actin-binding protein. We recently demonstrated that IP3K-A modulates a structural reorganization of dendritic spines through F-actin remodeling, which is required for synaptic plasticity and memory formation in brain. However, detailed functions of IP3K-A and its regulatory mechanisms involved in the neuronal cytoskeletal dynamics still remain unknown. In the present study, we identified tubulin as a candidate of IP3K-A-binding protein through proteomic screening. By various in vitro and in vivo approaches, we demonstrated that IP3K-A was a novel microtubule-associated protein (MAP), and the N terminus of IP3K-A was a critical region for direct binding to tubulin in dendritic shaft of hippocampal neurons. Moreover, PKA phosphorylated Ser-119 within IP3K-A, leading to a significant reduction of microtubule binding affinity. These results suggest that PKA-dependent phosphorylation and microtubule binding of IP3K-A are involved in its regulatory mechanism for activity-dependent neuronal events such as local calcium signaling and its synaptic targeting.

Developmental expression and subcellular distribution of synaptotagmin 11 in rat hippocampus.

Synaptotagmins are required for Ca(2+)-dependent membrane-trafficking in either neuronal synaptic vesicles or cellular membranes. Previous reports suggested that the synaptotagmin 11 (syt11) gene is involved in the development of schizophrenia based on the genomic analysis of patients. Parkin protein binds to the C2 domains of Syt11 which leads to the polyubiquitination of Syt11. However, where and how Syt11 performs its role in the brain is largely unknown. Here, we report that Syt11 is expressed mainly in the brain. In addition, exogenously expressed Syt11 in HEK293 cells can form higher molecular weight complex via its transmembrane domain. Also, Syt11 is targeted to both dendrite and axon compartments. Immunocytochemistry showed that Syt11 is juxtaposed to postsynaptic markers in both excitatory and inhibitory synapses. Both neuroligin 1 and 2, which are postsynaptic cell adhesion molecules and differentially induce excitatory and inhibitory presynapses, respectively, recruit Syt11 in neuron coculture. Immunogold electron microscopy analysis revealed that Syt11 exists mainly in presynaptic neurotransmitter vesicles and plasma membrane, and rarely in postsynaptic sites. These results suggest that Syt11 may contribute to the regulation of neurotransmitter release in the excitatory and inhibitory presynapses, and postsynapse-targeted membrane trafficking in dendrites.

Induction of CITED2 expression in the rat hippocampus following transient global ischemia

CITED2 is implicated in the modulating the activity of HIF-1 which is a major transcription factor involved in ischemia-related gene expression. Following transient forebrain ischemia, we found that CITED2 was induced in a subset of brain regions including dentate gyrus of the hippocampal formation and piriform cortex. Because CITED2 was not induced in cultured neurons exposed to oxygen-glucose deprivation, we concluded that hypoxia is not sufficient to trigger its induction.

Impaired Extinction of Learned Contextual Fear Memory in Early Growth Response 1 Knockout Mice

Inductive expression of early growth response 1 (Egr-1) in neurons is associated with many forms of neuronal activity. However, only a few Egr-1 target genes are known in the brain. The results of this study demonstrate that Egr-1 knockout (KO) mice display impaired contextual extinction learning and normal fear acquisition relative to wild-type (WT) control animals. Genome-wide microarray experiments revealed 368 differentially expressed genes in the hippocampus of Egr-1 WT exposed to different phases of a fear conditioning paradigm compared to gene expression profiles in the hippocampus of KO mice. Some of genes, such as serotonin receptor 2C (Htr2c), neuropeptide B (Npb), neuronal PAS domain protein 4 (Npas4), NPY receptor Y1 (Npy1r), fatty acid binding protein 7 (Fabp7), and neuropeptide Y (Npy) are known to regulate processing of fearful memories, and promoter analyses demonstrated that several of these genes contained Egr-1 binding sites. This study provides a useful list of potential Egr-1 target genes which may be regulated during fear memory processing.