title: Calpain 2 controls turnover of LFA-1 adhesions on migrating T lymphocytes.
abstract: The immune cells named T lymphocytes circulate around the body fulfilling their role in immunosurveillance by monitoring the tissues for injury or infection. To migrate from the blood into the tissues, they make use of the integrin LFA-1 which is exclusively expressed by immune cells. These highly motile cells attach and migrate on substrates expressing the LFA-1 ligand ICAM-1. The molecular events signaling LFA-1 activation and adhesion are now reasonably well identified, but the process of detaching LFA-1 adhesions is less understood. The cysteine protease calpain is involved in turnover of integrin-mediated adhesions in less motile cell types. In this study we have explored the involvement of calpain in turnover of LFA-1-mediated adhesions of T lymphocytes. Using live cell imaging and immunohistochemistry, we demonstrate that turnover of adhesions depends on the Ca2+-dependent enzyme, calpain 2. Inhibition of calpain activity by means of siRNA silencing or pharmacological inhibition results in inefficient disassembly of LFA-1 adhesions causing T lymphocyte elongation and shedding of LFA-1 clusters behind the migrating T lymphocytes. We show that calpain 2 is distributed throughout the T lymphocyte, but is most active at the trailing edge as detected by expression of its fluorescent substrate CMAC,t-BOC-Leu-Met. Extracellular Ca2+ entry is essential for the activity of calpain 2 that is constantly maintained as the T lymphocytes migrate. Use of T cells from a patient with mutation in ORAI1 revealed that the major calcium-release-activated-calcium channel is not the ion channel delivering the Ca2+. We propose a model whereby Ca2+ influx, potentially through stretch activated channels, is sufficient to activate calpain 2 at the trailing edge of a migrating T cell and this activity is essential for the turnover of LFA-1 adhesions.
authors.affiliation: Leukocyte Adhesion Laboratory, Cancer Research UK London Research Institute, London, United Kingdom.
authors.forename: Lena
authors.initials: L
authors.lastname: Svensson
authors.forename: Alison
authors.initials: A
authors.lastname: McDowall
authors.forename: Katherine M
authors.initials: KM
authors.lastname: Giles
authors.forename: Paula
authors.initials: P
authors.lastname: Stanley
authors.forename: Stefan
authors.initials: S
authors.lastname: Feske
authors.forename: Nancy
authors.initials: N
authors.lastname: Hogg
mesh_terms: D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism; D002154:Calpain / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002465:Cell Movement; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D016169:Lymphocyte Function-Associated Antigen-1 / Q000378:metabolism*; D018613:Microscopy, Confocal; D018715:Microscopy, Video; D009154:Mutation; D000071740:ORAI1 Protein; D034622:RNA Interference; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism*
chemical_list: D002121:Calcium Channel Blockers; D015220:Calcium Channels; D007093:Imidazoles; D016169:Lymphocyte Function-Associated Antigen-1; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; D002154:Calpain; C569870:CAPN2 protein, human; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; D002118:Calcium