Annual Report 2023
Division of Advanced Bioimaging
Kenichi Suzuki, Rinshi Kasai
Introduction
To understand how anticancer drugs work, clarifying how cancer-related gene products behave in cells is mandatory. Molecules in cells do not work synchronously but essentially work in a stochastic process theory (randomly). Individual molecules engage in interactions lasting less than a second, with the proportion of interacting molecules rarely exceeding 10%. Single-molecule imaging is a powerful tool to unravel such stochastic molecular events occurring in cells.
The Team and What We Do
Using single-molecule imaging at high-temporal and spatial resolution, we are trying to elucidate the basic principles of molecular events on cell membranes, which are key factors in the oncogenesis of cells. We have constructed a total internal reflection fluorescence microscopy system that enables us to simultaneously observe single molecules in three colors with a temporal resolution of 1 ms. We also constructed a 3D single-molecule and super-resolution observation system with a spatial resolution of 20 nm.
Research Activities
1) Elucidation of the activation mechanism of STING
The innate immune response molecule, STING, is important not only for infection immunity but also for cancer immune response. It has been reported that STING migrates from the endoplasmic reticulum to the Golgi upon DNA virus infection and that STING undergoes palmitoylation lipid modification, which is required for its activation. However, it was unclear how palmitoylated STING activates innate immune responses. We investigated the STING activation mechanism using the high-resolution single molecule and super-resolution microscopy system described above, and revealed the following:
A) Activated STING formed large clusters on the trans-Golgi network in living cells and the number of molecules within the clusters was quantified.
B) Palmitoylation of STING and cholesterol was essential for the formation of large STING clusters.
C) Phosphorylation of STING by the downstream signaling molecule TBK1 stabilized the binding of TBK1 to STING clusters.
2) Elucidation of the function of adhesive GPCR
We found that CELSR, an adhesive GPCR interacts with each other in the cell adhesion region to form an inversely parallel transdimeric structure. This was quite different from the typical E-cadherin structure.
Education
We accepted one graduate student from the United Graduate School of Agricultural Science of Gifu University and four undergraduate students from Hoshi University as observership visitors.
Future Prospects
We will continue to elucidate how oncogene products such as erbB and Ras facilitate signal transduction utilizing cell membrane structures. Furthermore, we will elucidate how this is altered by the expression of the mutants of these molecules. We will also elucidate the mechanisms of inter- and intra-cellular signal transduction involving the cancer-related protein Wnt, a ligand of GPCR. To address the question of how cancer progresses at the transcriptional level, we will investigate gene transcription and chromatin structure within the nucleus. Through these studies, we aim to contribute to the advancement of cancer research.
List of papers published in 2023
Journal
1. Kemmoku H, Takahashi K, Mukai K, Mori T, Hirosawa KM, Kiku F, Uchida Y, Kuchitsu Y, Nishioka Y, Sawa M, Kishimoto T, Tanaka K, Yokota Y, Arai H, Suzuki KGN, Taguchi T. Single-molecule localization microscopy reveals STING clustering at the trans-Golgi network through palmitoylation-dependent accumulation of cholesterol. Nature communications, 15:220, 2024
2. Kusumi A, Tsunoyama TA, Tang B, Hirosawa KM, Morone N, Fujiwara TK, Suzuki KGN. Cholesterol- and actin-centered view of the plasma membrane: updating the Singer-Nicolson fluid mosaic model to commemorate its 50th anniversary(†). Molecular biology of the cell, 34:pl1, 2023
3. Nishiguchi S, Kasai RS, Uchihashi T. Antiparallel dimer structure of CELSR cadherin in solution revealed by high-speed atomic force microscopy. Proceedings of the National Academy of Sciences of the United States of America, 120:e2302047120, 2023
4. Suzuki KGN, Komura N, Ando H. Recently developed glycosphingolipid probes and their dynamic behavior in cell plasma membranes as revealed by single-molecule imaging. Glycoconjugate journal, 40:305-314, 2023
5. Fujiwara TK, Takeuchi S, Kalay Z, Nagai Y, Tsunoyama TA, Kalkbrenner T, Iwasawa K, Ritchie KP, Suzuki KGN, Kusumi A. Development of ultrafast camera-based single fluorescent-molecule imaging for cell biology. The Journal of cell biology, 222:e202110160, 2023
6. Fujiwara TK, Tsunoyama TA, Takeuchi S, Kalay Z, Nagai Y, Kalkbrenner T, Nemoto YL, Chen LH, Shibata ACE, Iwasawa K, Ritchie KP, Suzuki KGN, Kusumi A. Ultrafast single-molecule imaging reveals focal adhesion nano-architecture and molecular dynamics. The Journal of cell biology, 222:e202110162, 2023
7. Isogami A, Higashi S, Okumura B, Shibata A, Hirosawa K, Suzuki K, Tsukiji S, Matsuura K, Ikeda M. Hierarchical supramolecular structure comprising reduction-responsive DNA microspheres and semi-artificial glycopeptide-based micro-asters. polymer journal , 55:1103–1107, 2023