α-Actinin-4/FSGS1 is required for Arp2/3-dependent actin assembly at the adherens junction.
We have developed an in vitro assay to study actin assembly at cadherin-enriched cell junctions. Using this assay, we demonstrate that cadherin-enriched junctions can polymerize new actin filaments but cannot capture preexisting filaments, suggesting a mechanism involving de novo synthesis. In agreement with this hypothesis, inhibition of Arp2/3-dependent nucleation abolished actin assembly at cell-cell junctions. Reconstitution biochemistry using the in vitro actin assembly assay identified α-actinin-4/focal segmental glomerulosclerosis 1 (FSGS1) as an essential factor. α-Actinin-4 specifically localized to sites of actin incorporation on purified membranes and at apical junctions in Madin-Darby canine kidney cells. Knockdown of α-actinin-4 decreased total junctional actin and inhibited actin assembly at the apical junction. Furthermore, a point mutation of α-actinin-4 (K255E) associated with FSGS failed to support actin assembly and acted as a dominant negative to disrupt actin dynamics at junctional complexes. These findings demonstrate that α-actinin-4 plays an important role in coupling actin nucleation to assembly at cadherin-based cell-cell adhesive contacts.
Department of Cell and Developmental Biology, University of Illinois, Urbana, IL 61801, USA.
Vivian W
VW
Tang
William M
WM
Brieher
D008841:Actin Cytoskeleton / Q000378:metabolism*; D051376:Actin-Related Protein 2-3 Complex / Q000502:physiology*; D000185:Actinin / Q000235:genetics / Q000502:physiology*; D000199:Actins / Q000032:analysis; D022005:Adherens Junctions / Q000378:metabolism*; D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002448:Cell Adhesion / Q000235:genetics; D002460:Cell Line; D004285:Dogs; D017354:Point Mutation; D058105:Polymerization
D051376:Actin-Related Protein 2-3 Complex; D000199:Actins; D015820:Cadherins; D000185:Actinin