Yoana Arroyo, Antonella Di Mambro, Yolanda Calle-Patino, Bela Wrench, John Gribben, Maria Esposito

Research output: Chapter in Book/Report/Conference proceedingConference contributionpeer-review


Background: Mixed lineage leukemia (MLL) is the most common leukemia in infant patients. This leukemia is due to chromosomal translocations at locus 11q23 and is associated with poor clinical outcome. Novel, targeted therapeutic approaches are needed. Protein phosphatase 2A (PP2A) is a serine threonine phosphatase which regulates the phosphorylation of several kinases, including Erk, Akt, GSK3 which are fundamental for MLL cells' survival. PP2A is a trimeric protein complex in which a core dimer formed between the scaffold subunit and the catalytic subunit is associated with one of the many regulatory subunits that facilitate and direct the interaction of the trimer with substrate proteins. PP2A activity is regulated by interaction with PP2A inhibitor proteins (PIPs), as well as by post- translational modifications of PP2A complex components.
Aims: In this context, our aim is to understand how PP2A post-translational modifications affect the activity of PP2A on its downstream targets and to investigate whether PP2A re-activation, by preventing phosphorylation and activation of collateral pathways, might represent a valid therapeutic strategy for MLL.
Methods: 12 different leukemic cell lines and 8 primary samples from MLL patients were included in this study. We measured the PP2A phosphatase activity by a colorimetric assay using a synthetic phospho-peptide and malachite green reagent. The protein levels of PP2A, phospho-PP2A, demethyl-PP2A, LCMT-1, PME-1, Akt, phospho-Akt, Erk, and phospho-Erk were determined by western blot using specific antibodies.
Results: Compared to healthy bone marrow, we found an increase in the phosphorylation at Tyrosine 307 (Tyr307) and a decrease in the methylation at Leucine 309 (Leu309) of the catalytic subunit of PP2A in MLL samples. These changes in the post-translational modifications of the catalytic subunit correlated with a decrease in the phosphatase activity of PP2A. Methylation at Leu309 has been described as an absolute requirement for the binding of the regulatory subunit B55α to PP2A core. B55α was found up-regulated in MLL cell lines and primary samples compared to healthy bone marrow.The methylation of the catalytic subunit of PP2A is catalyzed by leucine carboxyl methyltransferase (LCMT-1), which we found down-regulated in MLL samples, whereas demethylation is catalyzed by the phosphatase methylesterase (PME-1) that was found up-regulated in MLL. In line with the inactivation of PP2A in the MLL samples, PP2A downstream targets such as Akt and Erk were found hyper-phosphorylated in MLL samples. Summary/Conclusion: Our results suggest that PP2A phosphatase activity inhibition might sustain in MLL samples a persistent serine/threonine phosphorylation of PP2A substrates, such as Akt and Erk that mediate pro-survival and anti- apoptotic signals. This inhibition might be explained, at least in part, by phosphorylation and demethylation of PP2A catalytic subunit. A better understanding of the mechanisms that regulate PP2A activity could provide new strategies to rescue PP2A phosphatase activity and target fundamental mechanisms of leukemic survival and chemotherapy resistance.
Original languageEnglish
Title of host publicationHemaSphere
PublisherWolter Kluwer Health, Inc
Number of pages1
Publication statusAccepted/In press - 30 Apr 2019

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