The Mechanism of CD47-Dependent Killing of T Cells: Heterotrimeric Gi-Dependent Inhibition of Protein Kinase A

Abstract

CD47 has been implicated in both positive and negative regulation of T cells as well as in T cell death. To clarify the role of CD47 in T cell function, we have studied the mechanism of T cell death in response to CD47 ligands, including mAb 1F7, thrombospondin-1, and a CD47 agonist peptide derived from it. CD47−/− Jurkat T cells (JINB8) were resistant to killing by all three ligands, indicating the essential role of CD47. Primary human T cells were also killed by CD47 ligands, but only after activation with anti-CD3. CD47-mediated cell death occurred without active caspases, DNA fragmentation, or Bcl-2 degradation. Pretreatment of Jurkat and primary T cells with pertussis toxin (PTX) prevented CD47-mediated death, indicating the involvement of Giα. Pretreatment of T cells with 8-bromo cAMP, forskolin, or 3-isobutyl-1-methylxanthine prevented the CD47-mediated apoptosis, and 1F7 dramatically reduced intracellular cAMP levels, an effect reversed with PTX. H89 and protein kinase A (PKA) inhibitor peptide, a specific PKA inhibitor, prevented rescue of T cells by PTX, 8-bromo cAMP, and forskolin, indicating a direct role for one or more PKA substrates. Thus, CD47-mediated killing of activated T cells occurs by a novel pathway involving regulation of cAMP levels by heterotrimeric Giα with subsequent effects mediated by PKA.

Human Lymphocytes Interact Directly with CD47 through a Novel Member of the Signal Regulatory Protein (SIRP) Family

“…5, b and c). The CD47 mAb MCA911-coated beads show that not all CD47 mAb are capable of inducing apoptosis as has been previously described (40). Abs that cross-linked the CD47-associated integrin CD51/61 had no effect on apoptosis, indicating a lack of direct involvement by CD47-associating integrins.…”

Interactions between CD47 and Thrombospondin Reduce Inflammation

“…Members of the TSP family have been described as natural ligands for CD47, and TSP-1 has been shown to induce T cell apoptosis (22,42). TSP-2 has previously been shown to be highly up-regulated in inflamed skin 24 h after oxazolone challenge and is found prominently in blood vessels and dermal cells (27).…”

Cholesterol-independent Interactions with CD47 Enhance αvβ3 Avidity

“…Depletion of membrane cholesterol was shown to destabilize the ␣ v ␤ 3 -CD47-G i complex in C32 melanoma and ovarian carcinoma cells, suggesting that intramembrane contacts controlling its localization to rafts may promote or even be required for complex assembly and G i -coupled signaling (9). CD47-mediated signaling also occurs in Jurkat T cells, where treatment with certain anti-CD47 antibodies promotes apoptosis via a G i -mediated pathway involving cyclic AMP regulation of cAMP-dependent kinase (11). Ligation of CD47 with other antibodies has been shown to have a synergistic effect on T cell activation that required the MMS domain of CD47 for both the T cell response and for raft localization (10).…”