Identification of the PAK4 interactome reveals PAK4 phosphorylation of N-WASP and promotion of Arp2/3-dependent actin polymerization

Oncotarget
Miao ZhaoStaffan Strömblad

Abstract

p21-activated kinase 4 (PAK4) regulates cell proliferation, apoptosis, cell motility and F-actin remodeling, but the PAK4 interactome has not been systematically analyzed. Here, we comprehensively characterized the human PAK4 interactome by iTRAQ quantitative mass spectrometry of PAK4-immunoprecipitations. Consistent with its multiple reported functions, the PAK4 interactome was enriched in diverse protein networks, including the 14-3-3, proteasome, replication fork, CCT and Arp2/3 complexes. Because PAK4 co-immunoprecipitated most subunits of the Arp2/3 complex, we hypothesized that PAK4 may play a role in Arp2/3 dependent actin regulation. Indeed, we found that PAK4 interacts with and phosphorylates the nucleation promoting factor N-WASP at Ser484/Ser485 and promotes Arp2/3-dependent actin polymerization in vitro. Also, PAK4 ablation in vivo reduced N-WASP Ser484/Ser485 phosphorylation and altered the cellular balance between G- and F-actin as well as the actin organization. By presenting the PAK4 interactome, we here provide a powerful resource for further investigations and as proof of principle, we also indicate a novel mechanism by which PAK4 regulates actin cytoskeleton remodeling.

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Citations

Jun 19, 2019·The Journal of Cell Biology·John G LockMartin J Humphries
Jan 19, 2021·The Journal of Biological Chemistry·Ave EesmaaMart Saarma
Jun 26, 2020·Journal of Structural Biology·Ashwin K ChettyTitus J Boggon

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Datasets Mentioned

BETA
PXD004086

Methods Mentioned

BETA
GTPase
immunoprecipitation
pull-down
co-IP
protein assay
pulldowns
pull down
transfection
immunoprecipitations

Software Mentioned

Proteome Discoverer
Graphpad Prism
DAVID
AutoAnnotate
SequestHT
NIS
PhosphoSitePlus
BiNGO
Cytoscape

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