Premium
Phosphorylation of α‐actinin‐4 (ACTN4) at serine 159 mimics the biochemical and cellular effects of human kidney disease mutations
Author(s) -
Feng Di,
Krishnan Ramaswamy,
Stella Lorenzo,
Sche Monica,
Hartigan Chrisina R,
Pollak Martin R.
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.771.1
Subject(s) - phosphorylation , actin , mutant , actin cytoskeleton , microbiology and biotechnology , biology , chemistry , cytoskeleton , gene , biochemistry , cell
α‐actinin‐4 gene (ACTN4) mutations cause a form of kidney injury called familial focal segmental glomerulosclerosis (FSGS) in humans. Individuals who carry disease‐causing ACTN4 mutations initially present with proteinuria due to podocyte injury, and many eventually develop end stage kidney disease. The disease‐causing ACTN4 mutations are exclusively located within the actin‐binding domain (ABD) of the encoded protein and result in increasing actin binding. We hypothesized that post‐translational modification of ACTN4 leads to similar biochemical and cellular changes observed in the disease‐causing ACTN4 mutant protein. Using mass spectrometry, we detected ACTN4 is phosphorylated at the serine 159 site (S159) in cultured human podocytes. By molecular dynamics simulations, we found that phosphorylated S159 causes a relative rotation of CH1 and CH2 domains within the ABD of ACTN4, potentially altering the actin binding interface. Supporting this is our observation that phosphomimetic S159D ACTN4 protein (which mimics the effect of phosphorylation at S159) increased F‐actin bundling activity in comparison with WT ACTN4, similar to the effect of kidney disease‐causing mutant K255E ACTN4 in vivo. Furthermore, when expressed in cultured human podocytes, phosphomimetic S159D ACTN4 led to similar changes as seen in the K255E mutant ACTN4: more prominent formation of actin stress fibers and significantly higher contractile force measured by traction force microscopy. We conclude that phosphorylation of ACTN4 at S159 induces characteristics similar to the kidney disease‐causing ACTN4 mutant both in vitro and in vivo. These data provide evidence that phosphorylation of ACTN4 plays an important role in regulating the cytoskeletal dynamics and function of podocytes. Support or Funding Information R37DK059588 and T32DK007199