Cytoplasmic anchorage of L-selectin controls leukocyte capture and rollingby increasing the mechanical stability of the selectin tether

L-selectin is a leukocyte lectin that mediates leukocyte capture and rollin g in the vasculature. The cytoplasmic domain of L-selectin has been shown t o regulate leukocyte rolling. In this study, the regulatory mechanisms by w hich this domain controls L-selectin adhesiveness were investigated. We rep ort that an L-selectin mutant generated by truncation of the COOH-terminal 11 residues of L-selectin tail, which impairs association with the cytoskel etal protein alpha -actinin, could capture leukocytes to glycoprotein L-sel ectin ligands under physiological shear flow. However, the conversion of in itial tethers into rolling was impaired by this partial tail truncation, an d was completely abolished by a further four-residue truncation of the L-se lectin tail. Physical anchorage of both cell-free tail-truncated mutants wi thin a substrate fully rescued their adhesive deficiencies. Microkinetic an alysis of full-length and truncated L-selectin-mediated rolling at millisec ond temporal resolution suggests that the lifetime of unstressed L-selectin tether's is unaffected by cytoplasmic tail truncation. However, cytoskelet al anchorage of L-selectin stabilizes the selectin tether by reducing the s ensitivity of its dissociation rate to increasing shear forces. Low force s ensitivity (reactive compliance) of tether lifetime is crucial for selectin s to mediate leukocyte rolling under physiological shear stresses. This is the first demonstration that reduced reactive compliance of L-selectin teth ers is regulated by cytoskeletal anchorage, in addition to intrinsic mechan ical properties of the selectin-carbohydrate bond.