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.