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Koichi Takimoto - Research
Research Interests
Neurons, cardiac myocytes and other excitable cells produce various patterns of electrical activity. I am interested in understanding how this diversity is generated and altered under physiological and pathological conditions. This diversity is primarily determined by the assortment of channel genes expressed by each cell. In particular, K+ channels are major determinants of excitability, and are encoded by a large number of genes. Therefore, the research in our laboratory is aimed at understanding cell type-specific regulation of K+ channel gene expression and the roles of other proteins that influence the expression and function of these channels.
Our current research focuses on Kv4-family channels and their associating auxiliary subunits. Molecular, biochemical and electrophysiological techniques, as well as autofluorescent protein-based imaging, are being used for these studies. Ongoing and developing projects are:
1. Region-selective and disease-associated regulation of Kv4 and KChIP2 gene expression in the heart. The size of the transient outward current (Ito) is regionally different within the heart tissue. For example, Ito is larger in epicardial or outside layer and smaller in endocardial or inside layer across the left ventricular wall. This transmural gradient of Ito is generated by differential expression of Kv4.2 gene in rodents and KChIP2 in large animals including human. The expression of Ito and these channel subunits are also decreased in failing heart and hypertrophied myocardium. To identify transcriptional regulatory mechanisms for the region-selective and disease-associated regulation of Kv4.2 and KChIP2 genes, we are examining cellular signaling mechanisms and promoters for these genes. We also plan to generate transgenic animals for testing the identified promoters for their ability to drive the region-selective expression.
Left: The level of Kv4.2 mRNA, but not other Kv channel messages, differs across the left ventricular wall.
Right: Hypertrophy-inducing stimuli (aortic banding, an animal model of hypertention, in vivo, and the a-adrenergic agonist phenylephrine in vivo) markedly reduces KChIP2 mRNA expression.
2. Functional roles of the newly identified DPPY in dorsal root ganglion neurons.
The newly identified dipeptidyl peptidase homologue, DPPX (DPP6), tightly binds to Kv4 channels to control their expression and gating. We have identified the novel DPPX homologue, namely DPPY (DPP10). This gene product specifically interacts with Kv4 proteins and is highly expressed in dorsal root ganglia. Using heterologous expression systems and recombinant proteins, we are determining molecular intereactions that confer Kv4-DPPX/Y complex formation and gating effects. Also, the functional roles of DPPY in dorsal root ganglion sensory neurons are being investigated.
Left: Crystal structures of related proteins (DPPIV for DPPX/Y and NCS-1 for KChIP) were used to construct a predicted organization of DPPX/Y-Kv4 channel-KChIP complex.
Right: RT-PCR analysis indicates that DPPY is highly expressed in DRGs (L4 and L5).
3. Expression and roles of Kv4-KChIP complexes in uterine myometrium Regulation of uterine myometrial membrane excitability is essential for quiescence during pregnancy and the induction of slow-wave contraction at term. We identified dramatic changes in the expression of several Kv channel subunit mRNAs. We are planning to determine the roles of each Kv channel in maintenance of quiescence and slow-wave contraction.
