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OCRL phosphatase controls actin reorganization during human and mouse platelet spreading

Background: Phosphatidylinositol-4, 5-bisphosphate [PI(4, 5)P2] is a phosphoinositide found at the plasma membrane which regulates actin reorganization. Lowe oculocerebrorenal syndrome protein (OCRL) dephosphorylates PI(4, 5)P2 and its mutations cause Lowe syndrome which is characterized by congenital cataracts, central hypotonia, and renal proximal tubular dysfunction. Aberrant primary hemostasis was shown in some Lowe syndrome patients. Platelets (PLT) recognize the site of injury and undergo extensive actin reorganization for their full activation. Aims: We tested if the inhibition or deficiency of OCRL leads to impaired PLT function due to changes in the actin cytoskeleton. Methods: Human PLTs with pharmacologically inhibited OCRL and mouse platelets deficient in OCRL were examined for their ability to spread on fibrinogen. In addition, the effect of the OCRL inhibition was tested in human PLT for the phosphorylation status of several signaling pathways after thrombin activation. Results: Human OCRL-inhibited PLTs failed to fully spread on fibrinogen and contained more actin nodules in all tested time points of activation compared to the untreated PLTs. Actin nodules in OCRL inhibited PLTs colocalized with vinculin, ARP2/3, as well as with phosphotyrosine suggesting dysregulation of the cytoskeleton remodeling. When activated with thrombin, OCRL-inhibited PLTs showed reduced phosphorylation of MLC, but not of p38 or ERK. Impaired spreading on fibrinogen was further confirmed with OCRL deficient mouse PLTs that exhibited increased filopodia when compared to wild type PLTs. Conclusions: Our data suggest that OCRL might control actin cytoskeleton rearrangements during platelet activation and thus their proper function which could be mediated by reduced MLC phosphorylation.

OCRL, PI(4, 5)P2, platelets, Lowe syndrome

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