CK1α protects WAVE from degradation to regulate cell shape and motility in the immune response.

Alexander Hirschhäuser,Marianne Van Cann,Sven Bogdan

doi:10.1242/jcs.258891

Alexander Hirschhäuser, Marianne Van Cann + Show 1 more

Open Access

https://doi.org/10.1242/jcs.258891

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Abstract

ABSTRACTThe WAVE regulatory complex (WRC) is the main activator of the Arp2/3 complex, promoting lamellipodial protrusions in migrating cells. The WRC is basally inactive but can be activated by Rac1 and phospholipids, and through phosphorylation. However, the in vivo relevance of the phosphorylation of WAVE proteins remains largely unknown. Here, we identified casein kinase I alpha (CK1α) as a regulator of WAVE, thereby controlling cell shape and cell motility in Drosophila macrophages. CK1α binds and phosphorylates WAVE in vitro. Phosphorylation of WAVE by CK1α appears not to be required for activation but, rather, regulates its stability. Pharmacologic inhibition of CK1α promotes ubiquitin-dependent degradation of WAVE. Consistently, loss of Ck1α but not ck2 function phenocopies the depletion of WAVE. Phosphorylation-deficient mutations in the CK1α consensus sequences within the VCA domain of WAVE can neither rescue mutant lethality nor lamellipodium defects. By contrast, phosphomimetic mutations rescue all cellular and developmental defects. Finally, RNAi-mediated suppression of 26S proteasome or E3 ligase complexes substantially rescues lamellipodia defects in CK1α-depleted macrophages. Therefore, we conclude that basal phosphorylation of WAVE by CK1α protects it from premature ubiquitin-dependent degradation, thus promoting WAVE function in vivo. This article has an associated First Person interview with the first author of the paper.

Highlights

Cell shape changes require a dynamic remodeling of the actin cytoskeleton
We identified the casein kinase 1α gene as a candidate that most strongly affected lamellipodia formation and phenocopied wave depleted cells, characterized by a prominently reduced circularity index (Figure 1B-D; quantification in 1I)
A previous study suggests that the casein kinase 2 (CK2) phosphorylates mammalian WAVE2 on Ser482, 484, 488, 489, and 497 within the acidic domain of the VCA region which promotes Arp2/3 complex activity in vitro (Pocha and Cory, 2009)

Summary

Introduction

Cell shape changes require a dynamic remodeling of the actin cytoskeleton. The WASP family verprolin homologous protein (WAVE) is a central Arp2/3 regulator driving lamellipodial protrusions and cell migration in most eukaryotic cells. Several studies have shown that phosphorylation plays an important role in regulating WRC-Arp2/3-mediated actin filament branching and lamellipodia formation (Mendoza, 2013). A previous in vitro study identified multiple functional phosphorylation events within the acidic VCA domain of mammalian WAVE2 by the casein kinase 2 (CK2) that are required for its activity (Pocha and Cory, 2009). A more recent in vivo study confirmed that the C-terminal acidic domain of the Dictyostelium WAVE is basally phosphorylated at four phosphorylation sites by CK2 and suggested that a regulated dephosphorylation of a fraction of the cellular SCAR/WAVE pool is a key step in its activation during pseudopod dynamics (Ura et al, 2012). Loss- and gain-of-function analysis and pharmacological inhibition of CK1 further suggest that basal phosphorylation of VCA domain by CK1 is critical for WAVE stability rather than its activity in vivo

Results

Discussion

Experimental procedures