Sphingosine analogue drug FTY720 targets I2PP2A/SET and mediates lung tumour suppression via activation of PP2A-RIPK1-dependent necroptosis.
Bottom Line: Mechanistically, targeting I2PP2A/SET by FTY720 mediated PP2A/RIPK1-dependent programmed necrosis (necroptosis), but not by apoptosis.The RIPK1 inhibitor necrostatin and knockdown or genetic loss of RIPK1 prevented growth inhibition by FTY720.Expression of WT- or death-domain-deleted (DDD)-RIPK1, but not the kinase-domain-deleted (KDD)-RIPK1, restored FTY720-mediated necroptosis in RIPK1(-/-) MEFs.
Affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, SC, USA.Show MeSH
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Mentions: Because I2PP2A/SET-FTY720 binding activated PP2A, we explored the role of PP2A on RIPK1-induced necroptosis. Ectopic PP2Ac (catalytic domain) expression alone or in combination with FTY720 induced necroptosis, however, siRNA-mediated knockdown of RIPK1 prevented necroptosis in response to PP2Ac (Fig 9A). Induction of PP2A-dependent necroptosis by FTY720 was also detected in H157 (K-Ras mutant) and H827 (EGFR mutant) human lung cancer cells, in which inhibition of RIPK1 or PP2A using necrostastin or OA and knockdown of RIPK1 or PP2Ac prevented FTY720-mediated cell death compared to controls (Supporting Information Fig S12A–C). Interestingly, ectopic PP2Ac had no effect on FTY720-mediated cell death in RIPK1−/− MEFs compared to vector-transfected controls (Supporting Information Fig S13A). Interestingly, siRNA-mediated knockdown of RIPK3, a known inducer of necroptosis via MKLN activation (Sun et al, 2012), or Drp1, a downstream target of PGAM5, which is a recently identified necroptosis inducer (Wang et al, 2012), did not prevent FTY720-mediated A549 cell death (Supporting Information Fig S13B–D). In fact, knockdown of RIPK3 increased FTY720-mediated LDH release around twofold compared to controls (Supporting Information Fig S13D). Thus, these data indicate that PP2A activation plays an upstream role in selective RIPK1-dependent necroptosis by FTY720 in these cells.
Affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, SC, USA.