Two-Way Adaptor

Science Signaling  20 May 2008:
Vol. 1, Issue 20, pp. ec190
DOI: 10.1126/stke.120ec190

Integrins are dimeric proteins that bind the extracellular matrix (ECM), and signaling by these receptors is bidirectional. "Outside-in" signaling induces integrin clustering to affect adhesion-regulated cell behaviors such as migration and shape change. "Inside-out" signaling affects the binding properties of integrins by controlling their activation state. The cytoplasmic tail of the β subunit of integrins has been implicated in inside-out signaling, whereas cytoplasmic tails of both the α and β subunits are involved in outside-in signaling. Montanez et al. identify the focal adhesion adaptor protein Kindlin-2 as important for both types of integrin signaling. Mice homozygous for a null allele of kindlin-2 exhibited early lethality coincident with implantation, which suggested an adhesion defect. To examine the adhesion defect, the authors cultured embryonic stem cells (ESCs) on a feeder monolayer of cells or on defined ECM substrates. The kindlin-2–/– ESCs exhibited defective adherence to the feeder layer as well as to the defined ECM substrates. The defective adherence did not appear to be due to decreased abundance of integrins. Rather, the kindlin-2–/– ESCs exhibited reduced binding of a fibronectin fragment and an antibody 9EG7 that recognizes activated integrins, suggesting that kindlin-2 contributes to integrin activation. If maintained in suspension culture, ESCs will form embryoid bodies, and the mutant ESCs formed severely abnormal embryoid bodies that resembled those formed from β1-integrin knockout ESCs. Furthermore, although the cells in the kindlin-2–/– embryoid bodies appeared to form normal cell-cell junctions and produced basement membrane proteins, 9EG7 failed to bind the endoderm and epiblast cells adjacent to the basement membrane, which is consistent with an integrin-activation defect. Glutathione S-transferase pull-down experiments showed that Kindlin-2 bound to the cytoplasmic tail of β1- and β3-integrins at a site previously identified as required for bidirectional signaling and distinct from the binding site of talin, which functions in integrin activation. When overexpressed in CHO (Chinese hamster ovary) cells, Kindlin-2 and the FERM domain of talin, which is the region that binds the β subunit of integrins, synergistically activated a heterologously expressed integrin. Endoderm and epiblast cells from the kindlin-2–/– embryoid bodies showed altered organization of the actin cytoskeleton. Endoderm cells derived from kindlin-2 null embryoid bodies failed to form focal adhesion, adhere, and spread when plated on fibronectin. Even when integrins were chemically activated by the addition of MnCl2, the isolated kindlin-2–/– endoderm cells failed to spread and recruit integrin-linked kinase to the few focal adhesions that formed, suggesting that outside-in signaling by integrins was also compromised in the absence of Kindlin-2. Thus, Kindlin-2 appears to be required for both integrin activation and outside-in integrin signaling.

E. Montanez, S. Ussar, M. Schifferer, M. Bösl, R. Zent, M. Moser, R. Fässler, Kindlin-2 controls bidirectional signaling of integrins. Genes Dev. 22, 1325-1330 (2008). [Abstract] [Full Text]