Abstract
Cell-cell adhesion is central to morphogenesis and maintenance of epithelial cell state. We previously identified 27 candidate cell-cell adhesion regulatory proteins (CCARPs) whose down-regulation disrupts epithelial cell-cell adhesion during collective migration. Using a protein interaction mapping strategy, we found that 18 CCARPs link to core components of adherens junctions or desmosomes. We further mapped linkages between the CCARPs and other known cell-cell adhesion proteins, including hits from recent screens uncovering novel components of E-cadherin adhesions. Mechanistic studies of one novel CCARP which links to multiple cell-cell adhesion proteins, the phosphatase DUSP23, revealed that it promotes dephosphorylation of β-catenin at Tyr 142 and enhances the interaction between α- and β-catenin. DUSP23 knockdown specifically diminished adhesion to E-cadherin without altering adhesion to fibronectin matrix proteins. Furthermore, DUSP23 knockdown produced “zipper-like” cell-cell adhesions, caused defects in transmission of polarization cues, and reduced coordination during collective migration. Thus, this study identifies multiple novel connections between proteins that regulate cell-cell interactions and provides evidence for a previously unrecognized role for DUSP23 in regulating E-cadherin adherens junctions through promoting the dephosphorylation of β-catenin.
Highlights
For detection of Tyr phosphorylated adhesion proteins, or to detect changes in the interactions between α- and β-catenin, cells treated with indicated siRNAs or pervanadate (1 mM) as a positive control were lysed in 500 μl mammalian cell lysis buffer (MCLB) + PhosSTOP (Roche) +pervanadate (1 mM) per two 35 mm wells on ice for 20 min
We utilized a comparative IP/MS analysis[14] to identify high-confidence protein interactions for each of the 27 proteins that we previously identified as candidate regulators of cell-cell adhesion
This led to the identification of linkages amongst the 27 proteins, as well as linkages to previously described cell-cell adhesion proteins
Summary
DUSP23 as a novel phosphatase for received: 17 September 2015 accepted: 10 May 2016 Published: 03 June 2016 β-catenin. Mechanistic studies of one novel CCARP which links to multiple cell-cell adhesion proteins, the phosphatase DUSP23, revealed that it promotes dephosphorylation of β-catenin at Tyr 142 and enhances the interaction between α- and β-catenin. This study identifies multiple novel connections between proteins that regulate cell-cell interactions and provides evidence for a previously unrecognized role for DUSP23 in regulating E-cadherin adherens junctions through promoting the dephosphorylation of β-catenin. The dense packing of desmosomal cadherins has led to speculation that the role of these adhesions is primarily for mechanical support[1]; recent studies have revealed more precise roles of desmosomes in regulating cell proliferation and differentiation[6,7,8] As such, both adherens and desmosomal junctions are sites of intercellular connections that coordinate the actions of numerous proteins to promote the homeostasis of epithelial tissues. This study provides a rich connectivity map amongst both known and novel cell-cell adhesion regulatory proteins
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