Intercellular communication is directly mediated by gap junction apparatus that forms conduits through adjacent two cell membranes in most multicellular organisms. Assembly of gap junction channels, termed gap junction plaques, mediates electrical and chemical coupling between cells, which is essential for various biological events such as development, inflammation, cell death, immune responses, and muscle contractions. We aim to understand the broad range of functional significance of gap junction channels in vivo. Cryo-EM is a powerful tool for high-resolution structure determination, and we will further integrate functional analysis and computational science to elucidate the gating mechanism of gap junction channels.

Large pore channels permeate ions and larger molecules such as nucleotides, amino acids, and peptides. Pannexins are categorized as large pore channels and have been known to be associated with inflammation and ischemia via ATP release through the channels. We are studying the gating mechanism of large pore channels embedded in phospholipids. We collaborate on functional analysis with electrophysiology and MD simulation to promote understanding of the channel functionality.

Navs generate the rapid upstroke of action potentials in nerve cell axons.
We focus on elucidating the molecular mechanisms for sodium selectivity and activity regulation, utilizing both of crystallography and electrophysiology.

Sample preparation is a specifically essential factor when focusing on high-resolution structure determination with single particle cryo-EM. Protein particles should be embedded in thin amorphous ice without denaturing and take various orientations. We are studying the effective techniques to keep membrane proteins intact structures, removing as free detergent micelles as possible. High-resolution structure determination has been successful for several membrane proteins using lipid nanodisc reconstitution approach, and we hope to move closer to the structure determinations in the native biomembrane environment.

To understand both the fast conduction and the high selectivity of water molecules through water channel (aquaporin), we focused on determining the atomic structure of aquaporin buried in the lipid membrane close to in vivo situation. In particular, out of the 13 water channels in the human body, we choose aquaporin-4 and aquaporin-11, which is expressed in the brains and the kidney, respectively. We often use cryo-electron microscope and electron crystallography to solve the 3D structure. Furthermore, to elucidate the function in vivo including the higher functional relation between the brain and water channels, we also use freeze fracture technique, optical microscope, and stopped-flow analysis as well as molecular biology methods.

G-protein coupled receptors (GPCRs) are membrane proteins to mediate signal transduction of extracellular stimulation such as light, smell, hormone, and neurotransmitters into the cytoplasm. The physiological function of GPCR is diverse and is associated with human diseases. We are interested in the structure and function of GPCRs which are functionally important, and possible drug targets. We have reported the cryo-EM structures of the MrgD-Gi complex, which are involved in itch and pain. Based on these, the characteristic ligand recognition and activation mechanism of MrgD have been demonstrated.