Cell Structure and Function
Online ISSN : 1347-3700
Print ISSN : 0386-7196
ISSN-L : 0386-7196
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Displaying 1-3 of 3 articles from this issue
  • Eriko Deguchi, Michiyuki Matsuda, Kenta Terai
    Article type: Reviews and Mini-reviews
    2025 Volume 50 Issue 1 Pages 1-14
    Published: 2025
    Released on J-STAGE: January 22, 2025
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    Live imaging techniques have revolutionized our understanding of paracrine signaling, a crucial form of cell-to-cell communication in biological processes. This review examines recent advances in visualizing and tracking paracrine factors through four key stages: secretion from producing cells, diffusion through extracellular space, binding to target cells, and activation of intracellular signaling within target cells. Paracrine factor secretion can be directly visualized by fluorescent protein tagging to ligand, or indirectly by visualizing the cleavage of the transmembrane pro-ligands or plasma membrane fusion of endosomes comprising the paracrine factors. Diffusion of paracrine factors has been studied using techniques such as fluorescence correlation spectroscopy (FCS), fluorescence recovery after photobleaching (FRAP), fluorescence decay after photoactivation (FDAP), and single-molecule tracking. Binding of paracrine factors to target cells has been visualized through various biosensors, including GPCR-activation-based (GRAB) sensors and Förster resonance energy transfer (FRET) probes for receptor tyrosine kinases. Finally, activation of intracellular signaling is monitored within the target cells by biosensors for second messengers, transcription factors, and so on. In addition to the imaging tools, the review also highlights emerging optogenetic and chemogenetic tools for triggering the release of paracrine factors, which is essential for associating the paracrine factor secretion to biological outcomes during the bioimaging of paracrine factor signaling.

    Key words: paracrine signaling, live imaging, biosensors, optogenetics, chemogenetics

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  • Zhang Weisheng, Jun Nakayama, Yukino Inomata, Shigeki Higashiyama, Tor ...
    Article type: Research Article
    2025 Volume 50 Issue 1 Pages 15-24
    Published: 2025
    Released on J-STAGE: February 07, 2025
    Advance online publication: December 18, 2024
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    Supplementary material

    Extracellular signal-regulated kinase (ERK) regulates multiple cellular functions through distinct activation patterns. Genetically encoded fluorescent probes are instrumental in dissecting the ERK activity dynamics in living cells. Here we modified a previously reported Förster resonance energy transfer (FRET) probe for ERK, EKAREN5 by replacing its mTurquoise2 and YPet sequences with mTurquoise-GL and a synonymous codon variant of YPet, respectively. The modified biosensor, EKAREN5-gl, showed an increased sensitivity to EGF-induced ERK activation responding to a very low dose (20 pg/ml) of EGF stimulation. We quantitatively characterized two FRET-based ERK probes, EKAREN5 and EKAREN5-gl, and a subcellular kinase translocation-based probe, ERK-KTR. We found the three biosensors differently respond to EGF stimulations with different intensity, duration, and latency. Furthermore, we investigated how the minimal EGF-induced ERK activation affects the downstream transcription in HeLa cells by comprehensive transcriptional analysis. We found the minimal ERK activation leads to a distinct transcriptional pattern from those induced by higher ERK activations. Our study highlights the significance of sensitive fluorescent probes to understand cellular signal dynamics and the role of minimal ERK activation in regulating transcription.

    Key words: fluorescent probe, ERK, FRET, KTR

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  • Yuta Yonekawa, Kazuki Oikawa, Boldbaatar Bayarkhuu, Kizuna Kobayashi, ...
    Article type: Research Article
    2025 Volume 50 Issue 1 Pages 25-39
    Published: 2025
    Released on J-STAGE: February 13, 2025
    Advance online publication: December 27, 2024
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    Supplementary material

    Membrane stiffness is essential for cell migration, tumorigenesis, and development; however, the physical properties of intracellular membrane are poorly characterized. In this study, we internalized 20 nm magnetic nanoparticles (MNPs) into MCF7 human breast cancer cells and applied a magnetic field. We investigated whether magnetic field could induce membrane damage of the early endosomes by analyzing the colocalization of MNPs with galectin 3 (Gal3), a cytosolic protein recruited to the lumen of damaged organelles. We first tried to apply magnetic field by electromagnet, and found a direct-current (DC) magnetic field for five minutes increased the colocalization of the MNPs with Gal3, suggesting that the magnetic field damaged the endosomal membrane. We used a neodymium magnet to apply longer and stronger static magnetic fields. The static magnetic field more than 50 mT for five minutes started to damage endosomes, while 100 mT was the most effective. Longer exposure or higher magnetic field strengths did not induce further membrane damage. We confirmed that a Gal3 positive compartment was also positive for the early endosome marker, EEA1, suggesting that the external magnetic field induced membrane damage in the early endosomes. Our results indicate that a static magnetic field can control the membrane damage in early endosomes using internalized MNPs.

    Key words: magnetic nanoparticles, endosomes, membrane damage, organelle

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