Filamin A regulates platelet shape change and contractile force generation via phosphorylation of the myosin light chain

Biochem J. 2024 Oct 17;481(20):1395-1410. doi: 10.1042/BCJ20240114.

Abstract

Platelets are critical mediators of hemostasis and thrombosis. Platelets circulate as discs in their resting form but change shape rapidly upon activation by vascular damage and/or soluble agonists such as thrombin. Platelet shape change is driven by a dynamic remodeling of the actin cytoskeleton. Actin filaments interact with the protein myosin, which is phosphorylated on the myosin light chain (MLC) upon platelet activation. Actin-myosin interactions trigger contraction of the actin cytoskeleton, which drives platelet spreading and contractile force generation. Filamin A (FLNA) is an actin cross-linking protein that stabilizes the attachment between subcortical actin filaments and the cell membrane. In addition, FLNA binds multiple proteins and serves as a critical intracellular signaling scaffold. Here, we used platelets from mice with a megakaryocyte/platelet-specific deletion of FLNA to investigate the role of FLNA in regulating platelet shape change. Relative to controls, FLNA-null platelets exhibited defects in stress fiber formation, contractile force generation, and MLC phosphorylation in response to thrombin stimulation. Blockade of Rho kinase (ROCK) and protein kinase C (PKC) with the inhibitors Y27632 and bisindolylmaleimide (BIM), respectively, also attenuated MLC phosphorylation; our data further indicate that ROCK and PKC promote MLC phosphorylation through independent pathways. Notably, the activity of both ROCK and PKC was diminished in the FLNA-deficient platelets. We conclude that FLNA regulates thrombin-induced MLC phosphorylation and platelet contraction, in a ROCK- and PKC-dependent manner.

Keywords: Filamin A; cytoskeleton; myosin; phosphorylation; platelets; protease-activated receptor signaling.

MeSH terms

  • Animals
  • Blood Platelets* / drug effects
  • Blood Platelets* / metabolism
  • Cell Shape / drug effects
  • Filamins* / metabolism
  • Mice
  • Mice, Knockout
  • Myosin Light Chains* / metabolism
  • Phosphorylation
  • Protein Kinase C / metabolism
  • Thrombin / metabolism
  • Thrombin / pharmacology
  • rho-Associated Kinases / metabolism

Substances

  • Filamins
  • Myosin Light Chains
  • FlnA protein, mouse
  • rho-Associated Kinases
  • Protein Kinase C
  • Thrombin