CD2AP localizes to discreet junctional puncta in polarized MDCK cells. (a) Single deconvolved optical section showing colocalization of CD2AP with E-cadherin and actin (phalloidin). (b) Single deconvolved optical z sections scanning from the apical (0 µm) to lateral (0.8 µm) junction. Yellow arrowheads point to colocalization of CD2AP puncta with E-cadherin and actin. Purple arrowheads point to colocalization of CD2AP puncta with E-cadherin in the absence of actin. White arrowheads point to puncta with E-cadherin alone. (c) Single deconvolved optical section showing colocalization of CD2AP with α-actinin-4, E-cadherin, and actin (yellow arrowheads). A subset of E-cadherin puncta has α-actinin-4 and actin without CD2AP (white arrowheads), whereas another subset of E-cadherin puncta has CD2AP and actin without α-actinin-4 (purple arrowheads). There is also a population of CD2AP not associating with α-actinin-4, E-cadherin, or actin (orange arrowhead). (d) Single deconvolved optical section showing colocalization of CD2AP with E-cadherin at latrunculin-resistant actin clusters (yellow arrowheads).