PHAGOCYTES , GRANULOCYTES , AND MYELOPOIESIS Minimal amounts of kindlin-3 suf fi ce for basal platelet and leukocyte functions in mice

Integrins are cell adhesion receptors that consist of a and b subunits. They anchor cells to the extracellular matrix and assemble large signaling hubs, with which they regulate essential cellular processes including cell adhesion, migration, proliferation, survival, and differentiation. A hallmark of integrins is their ability to reversibly switch between an active and an inactive conformation. In their inactive conformation, they have low affinity for ligands, whereas upon activation (inside-out signaling), the affinity increases, leading to ligand binding, integrin clustering, and, finally, signaling (outside-in signaling). Two principal protein families control integrin activity, talins (talin-1 and -2) and kindlins (kindlin-1, -2, -3). They both bind directly to b-integrin tails, cooperate during integrin inside-out and outside-in signaling, and stabilize the integrin-ligand interaction by inducing integrin clustering. A fast allosteric change and stable integrin-ligand interaction is particularly important for blood cells such as platelets and leukocytes, whose surface integrins continuously encounter ligands in plasma or on the vascular endothelium, respectively. Therefore, it is of fundamental importance for these cells to rapidly activate their integrins on demand—for example, during bleeding when platelet aggregation is needed to seal vascular injuries, or during infection when leukocytes need to extravasate and kill microbial invaders. Consequently, loss of talin-1 and kindlin-3, which are expressed in all hematopoietic cells, leads to severe bleeding and immunodeficiency disorders. Despite this central role of talin-1 andkindlin-3 in controlling integrin activity, it is unclear whether they are expressed and bind integrin tails in stoichiometric quantities, whether they are present in excess over integrins in hematopoietic cells andwhether a concentration threshold is required for them to enable integrin-mediated cellular processes to proceed normally. In the present study, we engineered several mouse strains expressing different kindlin-3 protein levels. This allowed testing whether different talin-1/kindlin-3 ratios influence integrin-mediated functions. We found that 5% of normal kindlin-3 levels in hematopoietic cells are sufficient for embryonic and postnatal development. However, upon exposure to stress, the low levels of kindlin-3 impair adhesive functions of platelets and neutrophils. We also found that kindlin-3 and talin-1 are expressed in stoichiometric quantities in both cell types. By contrast, there are twice as many b2-integrin molecules and only half the number of b3 integrins as talin-1/kindlin-3 in neutrophils and platelets, respectively. The implications of these findings are discussed.