Mechanisms of cyclic AMP/protein kinase A- and glucocorticoid-mediated apoptosis using S49 lymphoma cells as a model system

Abstract

Cyclic AMP/protein kinase A (cAMP/PKA) and glucocorticoids promote the death of many cell types, including cells of hematopoietic origin. In wild-type (WT) S49 T-lymphoma cells, signaling by cAMP and glucocorticoids converges on the induction of the proapoptotic B-cell lymphoma-family protein Bim to produce mitochondria-dependent apoptosis. Kin–, a clonal variant of WT S49 cells, lacks PKA catalytic (PKA-Cα) activity and is resistant to cAMP-mediated apoptosis. Using sorbitol density gradient fractionation, we show here that in kin– S49 cells PKA-Cα is not only depleted but the residual PKA-Cα mislocalizes to heavier cell fractions and is not phosphorylated at two conserved residues (Ser338 or Thr197). In WT S49 cells, PKA-regulatory subunit I (RI) and Bim coimmunoprecipitate upon treatment with cAMP analogs and forskolin (which increases endogenous cAMP concentrations). By contrast, in kin– cells, expression of PKA-RIα and Bim is prominently decreased, and increases in cAMP do not increase Bim expression. Even so, kin– cells undergo apoptosis in response to treatment with the glucocorticoid dexamethasone (Dex). In WT cells, glucorticoid-mediated apoptosis involves an increase in Bim, but in kin– cells, Dex-promoted cell death appears to occur by a caspase 3-independent apoptosis-inducing factor pathway. Thus, although cAMP/PKA-Cα and PKA-R1α/Bim mediate apoptotic cell death in WT S49 cells, kin– cells resist this response because of lower levels of PKA-Cα and PKA-RIα subunits as well as Bim. The findings for Dex-promoted apoptosis imply that these lymphoma cells have adapted to selective pressure that promotes cell death by altering canonical signaling pathways.

Protein kinase A activation by the anti-cancer drugs ABT-737 and thymoquinone is caspase-3-dependent and correlates with platelet inhibition and apoptosis

“…On the other hand, substantial evidence indicates that pro-apoptotic events of cAMP in nucleated cells are, at least in part, mediated by the PKA-dependent phosphorylation of protein targets. 61 , 62 Given that the effects of ABT-737, TQ, and PKA on nucleated cell apoptosis partially involve regulation of gene expression, it is difficult to compare these findings to the situation in platelets. For example, in Jurkat T-cells, TQ downregulated the expression of phosphodiesterases PDE1, PDE3, and PDE4 during 24 h of incubation, which increased cGMP levels and reduced cAMP concentrations.…”

cAMP-dependent activation of the Rac guanine exchange factor P-REX1 by type I protein kinase A (PKA) regulatory subunits

“…on the association between PKA regulatory subunits and putative signaling effectors. These include RII␤ interacting with G␣ i in response to the coincident activation of G i -and G s -coupled receptors, enhancing MAPK signaling (20), and RI␣ interacting with Bim (B-cell lymphoma family protein), contributing to mitochondria-dependent apoptosis in S49 cells, also involving PKA catalytic activity (21). Consistent with a model in which RI␣ interacts with P-REX1 and acquires an active conformation that promotes P-REX1 activity, we found that RI␣ mutants with alterations at their CNB-B domain actually exhibited a better interaction with P-REX1 than WT RI␣.…”

Protein kinase A determines platelet life span and survival by regulating apoptosis

“…PKA activity is regulated by intracellular concentration of cAMP, which is determined by the balance between synthesis and degradation by adenylate cyclase (12) and phosphodiesterases (13), respectively. Both proapoptotic and antiapoptotic effects of PKA were reported in nucleated cells (14,15). PKA is highly expressed, and PKA activity is strictly balanced in platelets (16,17).…”