Talin-1 and Kindlin-3 Regulate α4β1 Integrin-Mediated Adhesion Stabilization, but Not G Protein-Coupled Receptor-Induced Affinity Upregulation

Abstract

Chemokine/chemoattractant G protein-coupled receptors trigger an inside–out signaling network that rapidly activates integrins, a key step in inflammatory leukocyte recruitment. Integrins mediate leukocyte arrest and adhesion to endothelium through multivalent binding, and they transmit outside–in signals to stabilize adhesion and coordinate cell spreading and migration. In the present study, we used RNA interference in the U937 monocytic cell line to investigate the role of talin-1, kindlin-3, and α-actinin-1 in the fMLF- and SDF-1α–induced upregulation of α4β1 integrin affinity and consequent adhesive events. Affinity upregulation of α4β1 integrin was not impaired by small interfering RNA knockdown of talin-1, kindlin-3, or α-actinin-1. Only kindlin-3 knockdown increased flow-induced detachment from VCAM-1–coated surfaces in response to fluid flow, whereas knockdown of either talin-1 or kindlin-3 increased detachment from ICAM-1–coated surfaces. Biochemical analyses revealed that α4β1 expression was highly enriched in U937 cell microridges and murine lymphocyte microvilli. Kindlin-3 was present throughout the cell, whereas talin-1 was largely excluded from microridges/microvilli. The subcellular colocalization of α4β1 and kindlin-3 in microridges may explain why kindlin-3 rapidly associates with α4β1 after G protein-coupled receptor signaling and contributes to adhesion strengthening. Talin-1 contributed to α4β1-dependent chemotaxis, suggesting that it participates in a later stage of the leukocyte adhesion cascade when the leukocyte cytoskeleton undergoes dramatic rearrangement.

ADAP interactions with talin and kindlin promote platelet integrin αIIbβ3 activation and stable fibrinogen binding

“…31,46,47 Conceivably, any or all of these adaptations may be regulated by ADAP, talin, and kindlin-3. 35,48,49 In this context, ADAP's enhancement of irreversible fibrinogen binding to aIIbb3 in platelets may reflect an underlying promotion of integrin clustering by ADAP, as was observed for b2 integrins in lymphocytes. 50,51 The signaling networks operating within different cellular environments (eg, platelets and lymphocytes) are likely to use cell context-dependent mechanisms to determine how ADAP, talin, and kindlins access and regulate integrins.…”

Kindlin-3 regulates integrin activation and adhesion reinforcement of effector T cells

“…This clearly indicates that β2 integrins induce outside-in signals even in the absence of Kindlin-3 and before reaching a fully active conformation. In contrast, the majority of Kindlin-3-deficient T cells attached to VCAM-1, ICAM-1, or TNF-α-treated endothelial cells are unable to resist higher shear forces, indicating that strengthening of integrin interactions with these ligands requires further changes in α4β1 and αLβ2 that are mediated by 32). It is possible that these changes represent conformational changes of the integrin ectodomain induced by tension and resulting in the formation of catch bonds that increase bond lifetimes.…”

B-Raf Regulation of Integrin α4β1-mediated Resistance to Shear Stress through Changes in Cell Spreading and Cytoskeletal Association in T Cells

“…However, it is established that different integrin heterodimers expressed by the same cells can utilize distinct signaling components and downstream effectors. For example, paxillin binding to ␣4 integrin cytoplasmic domains is known to regulate cytoskeletal association and resistance to shear stress but not affinity of ␣4␤1, whereas paxillin does not bind to ␣L of LFA-1 (5,30,45).…”