Summary

The Phactr family of PP1-binding proteins is implicated in human diseases including Parkinson's, cancer and myocardial infarction. Each Phactr protein contains a conserved C-terminus required for interaction with PP1, and four G-actin binding RPEL motifs: an N-terminal motif, abutting a basic element, and a C-terminal triple RPEL repeat, which overlaps the PP1 binding sequences. The MRTF family of transcriptional coactivators are also RPEL proteins: their regulatory RPEL domains sense signal-induced changes in G-actin concentration, and thereby control MRTF subcellular localisation and activity. However, whether G-actin binding controls Phactr protein function, and its relation to signalling, has not been investigated. Here we show that Rho-actin signalling by serum stimulation induces nuclear accumulation of Phactr1, but not other Phactr family members. Actin binding by the three Phactr1 C-terminal RPEL motifs is required for Phactr1 cytoplasmic localisation in resting cells. Phactr1 nuclear accumulation is importin α-β-dependent. G-actin and importin α-β bind competitively to nuclear import signals associated with the N- and C-terminal RPEL motifs. All four motifs are required for the inhibition of serum-induced Phactr1 nuclear accumulation by elevated G-actin. G-actin and PP1 bind competitively to the Phactr1 C-terminal region, and Phactr1 C-terminal RPEL mutants that cannot bind G-actin induce aberrant actomyosin structures dependent on their nuclear accumulation and on PP1 binding. In CHL-1 melanoma cells, Phactr1 exhibits actin-regulated subcellular localisation and is required for stress fibre assembly, motility, and invasiveness. These data support a role for Phactr1 in actomyosin assembly and suggest that Phactr1 G-actin sensing allows its coordination with F-actin availability.