Identification and Characterization of an Alternatively Spliced Isoform of the Human Protein Phosphatase 2Aα Catalytic Subunit

• Published in: The Journal of biological chemistry 2012-02, Vol.287 (7), p.4853-4862
• Main Authors: Migueleti, Deivid L.S., Smetana, Juliana H.C., Nunes, Hugo F., Kobarg, Jörg, Zanchin, Nilson I.T.
• Format: Article
• Language: English
• Subjects: Alternative splicing; Alternative Splicing - physiology; Carboxylic Ester Hydrolases - genetics; Carboxylic Ester Hydrolases - metabolism; Gene expression; HEK293 Cells; Humans; Isoenzymes - biosynthesis; Isoenzymes - genetics; Jurkat Cells; Leukocytes, Mononuclear - cytology; Leukocytes, Mononuclear - enzymology; Models, Molecular; Protein O-Methyltransferase - genetics; Protein O-Methyltransferase - metabolism; Protein Phosphatase 2 - biosynthesis; Protein Phosphatase 2 - genetics; Protein-protein interactions; RNA, Messenger - biosynthesis; RNA, Messenger - genetics; Serine threonine protein phosphatase; Signal Transduction
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M111.283341

PP2A is the main serine/threonine-specific phosphatase in animal cells. The active phosphatase has been described as a holoenzyme consisting of a catalytic, a scaffolding, and a variable regulatory subunit, all encoded by multiple genes, allowing for the assembly of more than 70 different holoenzymes. The catalytic subunit can also interact with α4, TIPRL (TIP41, TOR signaling pathway regulator-like), the methyl-transferase LCMT-1, and the methyl-esterase PME-1. Here, we report that the gene encoding the catalytic subunit PP2Acα can generate two mRNA types, the standard mRNA and a shorter isoform, lacking exon 5, which we termed PP2Acα2. Higher levels of the PP2Acα2 mRNA, equivalent to the level of the longer PP2Acα mRNA, were detected in peripheral blood mononuclear cells that were left to rest for 24 h. After this time, the peripheral blood mononuclear cells are still viable and the PP2Acα2 mRNA decreases soon after they are transferred to culture medium, showing that generation of the shorter isoform depends on the incubation conditions. FLAG-tagged PP2Acα2 expressed in HEK293 is catalytically inactive. It displays a specific interaction profile with enhanced binding to the α4 regulatory subunit, but no binding to the scaffolding subunit and PME-1. Consistently, α4 out-competes PME-1 and LCMT-1 for binding to both PP2Acα isoforms in pulldown assays. Together with molecular modeling studies, this suggests that all three regulators share a common binding surface on the catalytic subunit. Our findings add important new insights into the complex mechanisms of PP2A regulation. Background: The catalytic subunit of the PP2A phosphatase interacts with structural and regulatory proteins and can be regulated post-translationally by phosphorylation and methylation. Results: A novel alternatively spliced isoform of the PP2A catalytic subunit has been identified. Conclusion: The alternatively spliced isoform shows a specific interaction profile and no phosphatase activity. Significance: The alternatively spliced isoform may represent a novel mechanism of PP2A regulation.