title: In Vitro and in Vivo Characterization of Molecular Interactions between Calmodulin, Ezrin/Radixin/Moesin, and L-selectin.
abstract: L-selectin is a cell adhesion molecule that tethers leukocytes to the luminal walls of venules during inflammation and enables them to roll under the force of blood flow. Clustering of L-selectin during rolling is thought to promote outside-in signals that lead to integrin activation and chemokine receptor expression, ultimately contributing to leukocyte arrest. Several studies have underscored the importance of the L-selectin cytoplasmic tail in functionally regulating adhesion and signaling. Interestingly, the L-selectin tail comprises only 17 amino acids, and yet it is thought to bind simultaneously to several proteins. For example, constitutive association of calmodulin (CaM) and ezrin/radixin/moesin (ERM) to L-selectin confers resistance to proteolysis and microvillar positioning, respectively. In this report we found that recombinant purified CaM and ERM bound non-competitively to the same tail of L-selectin. Furthermore, molecular modeling supported the possibility that CaM, L-selectin, and moesin could form a heterotrimeric complex. Finally, using fluorescence lifetime imaging microscopy to measure fluorescence resonance energy transfer, it was shown that CaM, L-selectin, and ERM could interact simultaneously in vivo. Moreover, L-selectin clustering promoted CaM/ERM interaction in cis (i.e. derived from neighboring L-selectin tails). These results highlight a novel intracellular event that occurs as a consequence of L-selectin clustering, which could participate in transducing signals that promote the transition from rolling to arrest.
authors.affiliation: Cardiovascular Science Unit, National Heart and Lung Institute, Imperial College London, Hammersmith Campus, London W12 0NN.
authors.forename: David J
authors.initials: DJ
authors.lastname: Killock
authors.forename: Maddy
authors.initials: M
authors.lastname: Parsons
authors.forename: Marouan
authors.initials: M
authors.lastname: Zarrouk
authors.forename: Simon M
authors.initials: SM
authors.lastname: Ameer-Beg
authors.forename: Anne J
authors.initials: AJ
authors.lastname: Ridley
authors.forename: Dorian O
authors.initials: DO
authors.lastname: Haskard
authors.forename: Marketa
authors.initials: M
authors.lastname: Zvelebil
authors.forename: Aleksandar
authors.initials: A
authors.lastname: Ivetic
mesh_terms: D000818:Animals; D019556:COS Cells; D002147:Calmodulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D019041:L-Selectin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D036904:Leukocyte Rolling / Q000502:physiology*; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008840:Microfilament Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D020298:U937 Cells; D000099149:Ezrin
chemical_list: D002147:Calmodulin; D003598:Cytoskeletal Proteins; D019041:L-Selectin; D008565:Membrane Proteins; D008840:Microfilament Proteins; D046912:Multiprotein Complexes; D011994:Recombinant Proteins; D000099149:Ezrin; C071058:moesin; C071776:radixin