Abstract
p21-activated kinase 4 (PAK4) is a serine/threonine kinase known to function in
cytoskeletal organisation. PAK4 is purported to localise to podosomes; transient
adhesive structures that can degrade the extracellular matrix to facilitate rapid myeloid
cell migration. As yet the functional significance of PAK4 at podosomes remains
unclear. Treatment of TGF-β-differentiated human monocytic leukaemia (THP-1) cells
with a PAK4-targeted ATP competitive inhibitor (PAK4i) significantly reduces
podosome formation and induces the formation of focal adhesions. This switch in
adhesion phenotype confers a diminution of matrix degradative ability and reduced
cell migration speed. Even more strikingly, primary human macrophages lose virtually
all podosomes following PAK4 inhibition, whilst PAK1 inhibition has no effect,
highlighting a specific and crucial role for PAK4 in macrophage podosome turn-over.
Complementary to our inhibitor studies, stable shRNA-mediated knockdown of PAK4
in THP-1 cells also causes a significant reduction in podosome number which cannot
be rescued by expression of kinase dead PAK4, supporting a kinase-dependent role
for PAK4 in podosomes. Concomitant with a loss of PAK4 expression, phosphorylation
of Akt is perturbed. Using immunoprecipitation and superresolution 3B analysis we
have identified a specific localisation for PAK4 in the podosome ring, nearer to the
actin core than other ring proteins vinculin, paxillin and talin. We now propose a novel
function for PAK4 kinase activity intersecting with the Akt pathway at the podosome
ring:core interface which is essential in the regulation of macrophage podosome turnover.
cytoskeletal organisation. PAK4 is purported to localise to podosomes; transient
adhesive structures that can degrade the extracellular matrix to facilitate rapid myeloid
cell migration. As yet the functional significance of PAK4 at podosomes remains
unclear. Treatment of TGF-β-differentiated human monocytic leukaemia (THP-1) cells
with a PAK4-targeted ATP competitive inhibitor (PAK4i) significantly reduces
podosome formation and induces the formation of focal adhesions. This switch in
adhesion phenotype confers a diminution of matrix degradative ability and reduced
cell migration speed. Even more strikingly, primary human macrophages lose virtually
all podosomes following PAK4 inhibition, whilst PAK1 inhibition has no effect,
highlighting a specific and crucial role for PAK4 in macrophage podosome turn-over.
Complementary to our inhibitor studies, stable shRNA-mediated knockdown of PAK4
in THP-1 cells also causes a significant reduction in podosome number which cannot
be rescued by expression of kinase dead PAK4, supporting a kinase-dependent role
for PAK4 in podosomes. Concomitant with a loss of PAK4 expression, phosphorylation
of Akt is perturbed. Using immunoprecipitation and superresolution 3B analysis we
have identified a specific localisation for PAK4 in the podosome ring, nearer to the
actin core than other ring proteins vinculin, paxillin and talin. We now propose a novel
function for PAK4 kinase activity intersecting with the Akt pathway at the podosome
ring:core interface which is essential in the regulation of macrophage podosome turnover.
| Original language | English |
|---|---|
| Pages (from-to) | 3385-3393.E6 |
| Journal | Cell Reports |
| Volume | 29 |
| Issue number | 11 |
| DOIs | |
| Publication status | Published - 10 Dec 2019 |
Keywords
- Cells, Cultured
- Disulfides/pharmacology
- Extracellular Matrix/metabolism
- Focal Adhesions/metabolism
- HEK293 Cells
- Humans
- Myeloid Cells/drug effects
- Naphthols/pharmacology
- Phosphorylation
- Podosomes/metabolism
- Protein Kinase Inhibitors/pharmacology
- Proto-Oncogene Proteins c-akt/metabolism
- Signal Transduction
- THP-1 Cells
- p21-Activated Kinases/antagonists & inhibitors
