title: Dissociation of SHP-1 from spinophilin during platelet activation exposes an inhibitory binding site for protein phosphatase-1 (PP1).
abstract: We have recently shown that a critical regulatory node in the platelet signaling network lies immediately downstream of platelet receptors for thrombin and TxA2. This node is comprised of a scaffold protein (spinophilin, SPL), a protein tyrosine phosphatase (SHP-1), and either of the two members of the Regulators of G protein Signaling family predominantly expressed in platelets (RGS10 or RGS18). The SPL/RGS/SHP-1 complex is present in resting platelets, dissociating when thrombin or TxA2, but not ADP or collagen, activate SHP-1 and release RGS10 and RGS18 to dampen signaling. Here we demonstrate an additional regulatory role for spinophilin, showing that dissociation of SHP-1 from spinophilin is followed by an increase in the binding of spinophilin to PP1, a serine/threonine phosphatase whose binding site maps to a region close to the SHP-1 binding site. The increase in PP1 binding to spinophilin is limited to platelet agonists that cause dissociation of the complex and is selective for the α and γ isoforms of PP1. Studies in cell culture show that SHP-1 and PP1 can compete for binding to spinophilin and that binding inhibits PP1 activity since over-expression of wild type spinophilin, but not spinophilin with a disabled PP1 binding site, causes an increase in the phosphorylation of myosin light chain, a well-characterized PP1 substrate. Collectively, these results indicate that in addition to regulating RGS protein availability in resting platelets, spinophilin can serve as a time-dependent, agonist- and isoform-selective regulator of PP1, inhibiting its activity when decay of the SPL/RGS/SHP-1 complex releases SHP-1 from spinophilin, exposing a binding site for PP1.
authors.affiliation: Department of Medicine and Pharmacology, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Peisong
authors.initials: P
authors.lastname: Ma
authors.affiliation: Department of Medicine and Pharmacology, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Darci C
authors.initials: DC
authors.lastname: Foote
authors.affiliation: Department of Medicine and Pharmacology, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Sinnamon
authors.affiliation: Department of Medicine and Pharmacology, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Lawrence F
authors.initials: LF
authors.lastname: Brass
mesh_terms: D000818:Animals; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism / Q000502:physiology*; D016466:CHO Cells; D003412:Cricetulus; D006801:Humans; D008840:Microfilament Proteins / Q000097:blood*; D009419:Nerve Tissue Proteins / Q000097:blood*; D010766:Phosphorylation; D015539:Platelet Activation / Q000502:physiology*; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6 / Q000097:blood*; D020710:RGS Proteins / Q000097:blood; D017476:Receptors, Neuropeptide Y / Q000097:blood*; D013917:Thrombin / Q000378:metabolism; D000099273:Neurabins
chemical_list: D008840:Microfilament Proteins; D009419:Nerve Tissue Proteins; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D020710:RGS Proteins; D017476:Receptors, Neuropeptide Y; D013917:Thrombin; D000099273:Neurabins; C109491:neuropeptide Y4 receptor; C509249:PTPN6 protein, human