Transmembrane Pickets Connect Cyto- and Pericellular Skeletons Forming Barriers to Receptor Engagement

Abstract

Phagocytic receptors must diffuse laterally to become activated upon clustering by multivalent targets. Receptor diffusion, however, can be obstructed by transmembrane proteins ("pickets") that are immobilized by interacting with the cortical cytoskeleton. The molecular identity of these pickets and their role in phagocytosis have not been defined. We used single-molecule tracking to study the interaction between Fcγ receptors and CD44, an abundant transmembrane protein capable of indirect association with F-actin, hence likely to serve as a picket. CD44 tethers reversibly to formin-induced actin filaments, curtailing receptor diffusion. Such linear filaments predominate in the trailing end of polarized macrophages, where receptor mobility was minimal. Conversely, receptors were most mobile at the leading edge, where Arp2/3-driven actin branching predominates. CD44 binds hyaluronan, anchoring a pericellular coat that also limits receptor displacement and obstructs access to phagocytic targets. Force must be applied to traverse the pericellular barrier, enabling receptors to engage their targets.

TNF-Stimulated Gene-6 Is a Key Regulator in Switching Stemness and Biological Properties of Mesenchymal Stem Cells

“…TSG‐6 is a potent modulator of HA binding to CD44 on lymphoid cell lines , which likely regulates leukocyte migration to the sites of inflammation. Interestingly, a recent study has demonstrated that the cell surface distribution of CD44 can restrict the movement of other receptors and influence the organization of the actin cytoskeleton . Our flow cytometry analyses revealed a reduction in the expression of CD44 on the surface of TSG‐6 −/− ‐MSCs compared with WT cells.…”

Inflamed synovial fluid induces a homeostatic response in bone marrow mononuclear cells in vitro: Implications for joint therapy

“…Expression of 27 gene targets was selected based on commonly used markers of inflammatory M1-like (CD86, TNF-α, IL-1α, IL-1β, IL-6, IDO1, STAT1, STAT4, CXCR4, and SDF-1), regulatory M2-like (CD206, IL-10, IL-4, IL-4r, PTGS2, IGF-1, TGF-β1, GPR86, EGR2, CXCL14, STAT3, and STAT6), and hematopoietic progenitors or naïve macrophages (MØ) (CD34, CD14, CD68, and CSF-1), or involved in cell adhesion and membrane reorganization (CD44). 7,57,62,63,[72][73][74] Cells recovered in 10 mM EDTA, were centrifuged (12 000 g; 10 minutes; 4°C) and the cell pellet placed in guanidinium chloride-phenol (Trizol, Life Technologies, 15 596 018, Carlsbad, CA). RNA was column-purified with on-column DNase digest (DirectZol RNA MicroPrep Kit, R2061, Zymo Research, Irvine, CA), quantified (Qubit 3.0 Fluorometer, 33216, ThermoFisher Scientific, Carlsbad, CA), and stored at −80°C.…”

Single-cell immuno-mechanics: rapid viscoelastic changes are a hall-mark of early leukocyte activation

“…Although the molecular details leading to changes in viscoelastic properties are yet to be understood, our observations that cell viscosity increases during leukocyte activation might imply that diffusive properties change during leukocyte activation. The formation of diffusional barriers could be one manifestation of the increase in viscosity that we observe on the whole cell level 69,70 . Diffusion and polarization of organelles such as mitochondria in T cells 71 might also be impacted by large viscous changes during leukocyte activation.…”