Direct visualization by FRET-FLIM of a putative mechanosome complex involving Src, Pyk2 and MBD2 in living MLO-Y4 cells

• Published in: PloS one Vol. 16; no. 12; p. e0261660
• Main Authors: Day, Richard N., Day, Kathleen H., Pavalko, Fredrick M.
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 23.12.2021; Public Library of Science (PLoS)
• Subjects: Anatomy & physiology; Animals; Biochemistry; Biological research; Biology; Biology and Life Sciences; Biology, Experimental; Biomedical materials; Bone cells; Cell adhesion molecules; Cell Line; Cell surface; Cytochemistry; Cytoplasm; DNA binding proteins; DNA-Binding Proteins - analysis; DNA-Binding Proteins - metabolism; Energy transfer; Extracellular matrix; Fluid dynamics; Fluid flow; Fluorescence Resonance Energy Transfer; Focal Adhesion Kinase 2 - analysis; Focal Adhesion Kinase 2 - metabolism; Focal Adhesions - metabolism; Gene expression; Kinases; Mechanical stimuli; Mechanoreceptors; Mechanotransduction, Cellular; Medicine and Health Sciences; Mice; Microscopy; Osteocytes; Osteocytes - cytology; Osteocytes - metabolism; Physical Sciences; Physiological aspects; Physiology; Protein tyrosine kinase; Proteins; PYK2 protein; Research and Analysis Methods; Shear stress; Signal transduction; Spatial discrimination; Spatial resolution; Src protein; src-Family Kinases - analysis; src-Family Kinases - metabolism; Visualization; United States; Indiana; United States > US; Indianapolis Indiana
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0261660

Earlier, we proposed the “mechanosome” concept as a testable model for understanding how mechanical stimuli detected by cell surface adhesion molecules are transmitted to modulate gene expression inside cells. Here, for the first time we document a putative mechanosome involving Src, Pyk2 and MBD2 in MLO-Y4 osteocytes with high spatial resolution using FRET-FLIM. Src-Pyk2 complexes were concentrated at the periphery of focal adhesions and the peri-nuclear region. Pyk2-MBD2 complexes were located primarily in the nucleus and peri-nuclear region. Lifetime measurements indicated that Src and MBD2 did not interact directly. Finally, mechanical stimulation by fluid flow induced apparent accumulation of Src-Pyk2 protein complexes in the peri-nuclear/nuclear region, consistent with the proposed behavior of a mechanosome in response to a mechanical stimulus.