期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：41
页码：12681-12686
DOI：10.1073/pnas.1516057112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceCyclic AMP, the first identified second messenger, regulates a wide array of cellular functions including apoptosis by activating protein kinase A (PKA) and, in turn, the phosphorylation of target proteins. The current study uses a variety of biochemical and functional analyses to assess wild-type S49 lymphoma cells and kin-, a clonal variant that lacks PKA. The results identify key alterations in the ability of kin- cells to process PKA and also define previously unidentified alterations in cAMP- and glucocorticoid-promoted killing of kin- S49 cells. The findings provide evidence for PKA-dependent pathway switching in cell death responses and have implications for therapeutic development in diseases with aberrant apoptosis. 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-C) activity and is resistant to cAMP-mediated apoptosis. Using sorbitol density gradient fractionation, we show here that in kin- S49 cells PKA-C is not only depleted but the residual PKA-C 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-RI 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-C and PKA-R1/Bim mediate apoptotic cell death in WT S49 cells, kin- cells resist this response because of lower levels of PKA-C and PKA-RI 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.
