Dynamic coupling of ALCAM to the actin cortex strengthens cell adhesion to CD6.
At the immunological synapse, the activated leukocyte cell adhesion molecule (ALCAM) on a dendritic cell (DC) and CD6 molecules on a T cell contribute to sustained DC-T-cell contacts. However, little is known about how ALCAM-CD6 bonds resist and adapt to mechanical stress. Here, we combine single-cell force spectroscopy (SCFS) with total-internal reflection fluorescence microscopy to examine ALCAM-CD6-mediated cell adhesion. The combination of cells expressing ALCAM constructs with certain cytoplasmic tail mutations and improved SCFS analysis processes reveal that the affinity of ALCAM-CD6 bonds is not influenced by the linking of the intracellular domains of ALCAM to the actin cortex. By contrast, the recruitment of ALCAM to adhesion sites and the propensity of ALCAM to anchor plasma membrane tethers depend on actin cytoskeletal interactions. Furthermore, linking ALCAM to the actin cortex through adaptor proteins stiffens the cortex and strengthens cell adhesion. We propose a framework for how ALCAMs contribute to DC-T-cell adhesion, stabilize DC-T-cell contacts and form a mechanical link between CD6 and the actin cortex to strengthen cell adhesion at the immunological synapse.
Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Geert Grooteplein 28, 6525 GA Nijmegen, The Netherlands.
Joost
J
Te Riet
Jonne
J
Helenius
Nico
N
Strohmeyer
Alessandra
A
Cambi
Carl G
CG
Figdor
Daniel J
DJ
Müller
D000199:Actins / Q000378:metabolism*; D015703:Antigens, CD / Q000378:metabolism*; D000945:Antigens, Differentiation, T-Lymphocyte / Q000378:metabolism*; D002448:Cell Adhesion / Q000502:physiology*; D015816:Cell Adhesion Molecules, Neuronal / Q000378:metabolism*; D003713:Dendritic Cells / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D005326:Fetal Proteins / Q000378:metabolism*; D006801:Humans; D020014:K562 Cells; D018625:Microscopy, Atomic Force; D008856:Microscopy, Fluorescence; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D020410:Activated-Leukocyte Cell Adhesion Molecule
D000199:Actins; D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D015816:Cell Adhesion Molecules, Neuronal; D005326:Fetal Proteins; C105613:ALCAM protein, human; D020410:Activated-Leukocyte Cell Adhesion Molecule; C062279:CD6 antigen
ALCAM; Atomic force microscopy; CD6; Cell adhesion; Immunological synapse; Membrane tethers; Single-cell force spectroscopy