Coactosin-Like 1 Antagonizes Cofilin to Promote Lamellipodial Protrusion at the Immune Synapse

Joanna Kim,Virginia S Shapiro,Michael J Shapiro,Pinar Gurel,Daniel D Billadeau,Puspa Thapa,Henry N Higgs,Adebowale O Bamidele,Karen E Hedin,Inari Kursula

doi:10.1371/journal.pone.0085090

Abstract

Actin depolymerizing factor-homology (ADF-H) family proteins regulate actin filament dynamics at multiple cellular locations. Herein, we have investigated the function of the ADF-H family member coactosin-like 1 (COTL1) in the regulation of actin dynamics at the T cell immune synapse (IS). We initially identified COTL1 in a genetic screen to identify novel regulators of T cell activation, and subsequently found that it associates with F-actin and localizes at the IS in response to TCR+CD28 stimulation. Live cell microscopy showed that depletion of COTL1 protein impaired T cell spreading in response to TCR ligation and abrogated lamellipodial protrusion at the T cell – B cell contact site, producing only a band of F-actin. Significantly, re-expression of wild type COTL1, but not a mutant deficient in F-actin binding could rescue these defects. In addition, COTL1 depletion reduced T cell migration. In vitro studies showed that COTL1 and cofilin compete with each other for binding to F-actin, and COTL1 protects F-actin from cofilin-mediated depolymerization. While depletion of cofilin enhanced F-actin assembly and lamellipodial protrusion at the IS, concurrent depletion of both COTL1 and cofilin restored lamellipodia formation. Taken together, our results suggest that COTL1 regulates lamellipodia dynamics in part by protecting F-actin from cofilin-mediated disassembly.

Highlights

The actin cytoskeleton participates in many cellular processes including immune synapse (IS) formation during T cell activation [1]
As the first gene identified by this screen, NFkB activating protein (NKAP) provided a proof-ofprinciple for the approach, we repeated the screen to identify additional molecules
Loss of coactosin-like 1 (COTL1) expression may be the cause of the T cell activation defect in JREM 474, implying that COTL1 is required for T cell activation

Summary

Introduction

The actin cytoskeleton participates in many cellular processes including immune synapse (IS) formation during T cell activation [1]. Cofilin, an actin depolymerizing factor-homology (ADF-H) family member severs ADP-F-actin via conformational changes in filament structure and depolymerizes aged filaments at the pointed ends [13] Together, this dynamic process of filament nucleation, severing and depolymerization synergize to produce a large pool of new actin barbed ends and free actin monomers at the leading edge that support and maintain lamellipodial protrusion. This dynamic process of filament nucleation, severing and depolymerization synergize to produce a large pool of new actin barbed ends and free actin monomers at the leading edge that support and maintain lamellipodial protrusion Based on this information, it might be expected that the actin severing and depolymerizing activity of cofilin would be required to promote or maintain lamellipodia formation, but depletion of cofilin results in expanded lamellipodial protrusion in several cell models [14,15,16] suggesting that in some cellular systems cofilin regulates actin filament dynamics by accelerating actin filament disassembly

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Conclusion