Calcium influx promotes PLEKHG4B localization to cell-cell junctions and regulates the integrity of junctional actin filaments

  • Mol Biol Cell. 2023 Dec 13:mbcE23050154. doi: 10.1091/mbc.E23-05-0154.
Komaki Ninomiya  1  2 ,  Kai Ohta  1 ,  Ukyo Kawasaki  1 ,  Shuhei Chiba  1 ,  Takanari Inoue  3 ,  Erina Kuranaga  2  4 ,  Kazumasa Ohashi  1  5 ,  Kensaku Mizuno  1  5
Affiliations
  • 1. Laboratory of Molecular and Cellular Biology, Graduate School of Life Sciences, Tohoku University, Aobayama, Sendai, Miyagi 980-8578, Japan.
  • 2. Laboratory for Histogenetic Dynamics, Graduate School of Life Sciences, Tohoku University, Aobayama, Sendai, Miyagi 980-8578, Japan.
  • 3. Department of Cell Biology and Center for Cell Dynamics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
  • 4. Laboratory for Histogenetic Dynamics, Graduate School and Faculty of Pharmaceutical Sciences, Kyoto University, Kyoto 606‑8304, Japan.
  • 5. Department of Chemistry, Graduate School of Science, Tohoku University, Aobayama, Sendai, Miyagi 980-8578, Japan.
PMID: 38088892 DOI: 10.1091/mbc.E23-05-0154
Abstract

PLEKHG4B is a Cdc42-targeting guanine-nucleotide exchange factor implicated in forming epithelial cell-cell junctions. Here we explored the mechanism regulating PLEKHG4B localization. PLEKHG4B localized to the basal membrane in normal CA2+ medium but accumulated at cell-cell junctions upon ionomycin treatment. Ionomycin-induced junctional localization of PLEKHG4B was suppressed upon disrupting its annexin-A2 (AnxA2)-binding ability. Thus, CA2+ influx and AnxA2 binding are crucial for PLEKHG4B localization to cell-cell junctions. Treatments with low CA2+ or BAPTA-AM (an intracellular CA2+ chelator) suppressed PLEKHG4B localization to the basal membrane. Mutations of the phosphoinositide-binding motif in the pleckstrin homology (PH) domain of PLEKHG4B or masking of membrane phosphatidylinositol-4,5-biphosphate [PI(4,5)P2] suppressed PLEKHG4B localization to the basal membrane, indicating that basal membrane localization of PLEKHG4B requires suitable intracellular CA2+ levels and PI(4,5)P2 binding of the PH domain. Activation of mechanosensitive ion channels (MSCs) promoted PLEKHG4B localization to cell-cell junctions, and their inhibition suppressed it. Moreover, similar to the PLEKHG4B knockdown phenotypes, inhibition of MSCs or treatment with BAPTA-AM disturbed the integrity of actin filaments at cell-cell junctions. Taken together, our results suggest that CA2+ influx plays crucial roles in PLEKHG4B localization to cell-cell junctions and the integrity of junctional actin organization, with MSCs contributing to this process. [Media: see text] [Media: see text].

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