Protein Phosphatase 2A Plays an Important Role in Stromal Cell-Derived Factor-1/CXC Chemokine Ligand 12-Mediated Migration and Adhesion of CD34+ Cells

Abstract

Migration of hemopoietic stem and progenitor cells (HSPC) is required for homing to bone marrow following transplantation. Therefore, it is critical to understand signals underlying directional movement of HSPC. Stromal cell-derived factor-1 (SDF-1)/CXCL12 is a potent chemoattractant for HSPC. In this study, we demonstrate that the serine-threonine protein phosphatase (PP)2A plays an important role in regulation of optimal level and duration of Akt/protein kinase B activation (a molecule important for efficient chemotaxis), in response to SDF-1. Inhibition of PP2A, using various pharmacological inhibitors of PP2A including okadaic acid (OA) as well as using genetic approaches including dominant-negative PP2A-catalytic subunit (PP2A-C) or PP2A-C small interfering RNA, in primary CD34+ cord blood (CB) cells led to reduced chemotaxis. This was associated with impairment in polarization and slower speed of movement in response to SDF-1. Concomitantly, SDF-1-induced Akt phosphorylation was robust and prolonged. Following SDF-1 stimulation, Akt and PP2A-C translocate to plasma membrane with enhanced association of PP2A-C with Akt observed at the plasma membrane. Inhibition of PI3K by low-dose LY294002 partially recovered chemotactic activity of cells pretreated with OA. In addition to chemotaxis, adhesion of CD34+ cells to fibronectin was impaired by OA pretreatment. Our study demonstrates PP2A plays an important role in chemotaxis and adhesion of CD34+ CB cells in response to SDF-1. CD34+ CB cells pretreated with OA showed impaired ability to repopulate NOD-SCID mice in vivo, suggesting physiological relevance of these observations.

Inactivation of Host Akt/Protein Kinase B Signaling by Bacterial Pore-forming Toxins

“…The composition and regulation of PP1 is similarly complex, with ␣, ␣ 2 , ␤, ␥ 1 , or ␥ 2 PP1 catalytic subunits being capable of interacting with Ͼ50 regulatory subunits (Cohen, 2002). Using siRNA, we attempted to verify a role for PP2A and PP1 in HlyAmediated dephosphorylation of Akt by knocking down expression of the PP2A and PP1 ␣-catalytic subunits, both of which have been previously implicated in the regulation of Akt Xu et al, 2003;Basu et al, 2007). However, siRNA-mediated knockdown of the ␣-catalytic subunits, either individually or together, had no effect on HlyA-mediated Akt dephosphorylation in our assays.…”

Human CD34 + /KDR + Cells Are Generated From Circulating CD34 + Cells After Immobilization on Activated Platelets

“…12 Adherent cells showed a clear signal for p-Akt ( Figure 2E-2G) as opposed to the nonperfused control cells ( Figure 2C). The presence of membrane-associated p-Akt at the contact sites of CD34 ϩ cells and ECs ( Figure 2F and 2G, different angles) again confirms that the CD34 ϩ cells are rapidly activated on firm adhesion.…”

Beyond Desensitization: Physiological Relevance of Arrestin-Dependent Signaling

“…Of these, the involvement of G proteinindependent signaling is most convincing for the AT 1A receptor, because cells will follow a gradient of the arrestin pathway-selective AT 1A receptor agonist Sar 1 -Ile 4 -Ile 8 , a response that is lost when arrestins are down-regulated. In addition to regulating cortical actin assembly through arrestin-dependent activation of ERK1/2 and a cofilin-LIM kinase-chronophin complex (DeFea, 2007), recent evidence suggests that other arrestin-regulated effectors, including the Src family kinase, Lyn (Cheung et al, 2009), PP2A (Basu et al, 2007), and the small GTPase, Ral (de Gorter et al, 2008;Li et al, 2009) are involved in chemokineinduced chemotaxis in vitro.…”