The protein serine-threonine phosphatase 2A (PP2A) family of enzymes participates in a wide spectrum of cellular functions including cell growth, differentiation, cell cycle progression, and apoptosis. Multiple regulatory mechanisms control the activity of PP2A in vivo. Preliminary findings from the applicant's laboratory have revealed a novel post-translational modification of the PP2A catalytic subunit (PP2Ac), namely monoubiquitination, which has recently been shown to play an essential role in directing the subcellular localization of target proteins. Consistent with these findings, a recent study from the applicant's collaborator has revealed defective ubiquitin-dependent regulation of PP2Ac as the underlying cause of Opitz Syndrome (OS), a disorder characterized by malformations of the ventral midline (Nat. Gen. 29:287, 2001). Mutations in the Midi gene, which are found in OS patients, lead to marked accumulation of microtubule-associated PP2Ac. This defect likely represents an impairment of the E3 ubiquitin ligase activity of the Midi protein that normally facilitates PP2Ac polyubiquitination and subsequent degradation. The specificity of Midi for PP2Ac is defined by its interaction with alpha4, a known interacting partner of PP2Ac that also has been linked to another OS-like condition known as FG syndrome. The applicant hypothesizes that Mid1/alpha4-dependent mono- and polyubiquitination of PP2Ac determine the enzyme's subcellular localization and degradation, respectively, thereby influencing phosphatase activity towards key signaling enzymes involved in the control of cell growth and proliferation including p70 S6 kinase and mitogen-activated protein kinases. To address this central hypothesis, the applicant proposes a series of complementary biochemical, immunological, and genetic investigations to elucidate the molecular mechanism, regulation, and function of PP2Ac ubiquitination. These studies will provide fundamental mechanistic insights into the pathobiochemistry of OS and FG syndrome, as well as the role of this posttranslational modification in the control of signal transduction pathways mediating cell growth and survival. Given the importance of PP2A in the maintenance of cell homeostasis, the proposed studies will undoubtedly identify novel targets to manipulate phosphatase activity within specific signaling pathways, and potential therapeutics for diseases affecting normal cell growth and survival.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
Project #
Application #
Study Section
Cancer Molecular Pathobiology Study Section (CAMP)
Program Officer
Blondel, Olivier
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Vanderbilt University Medical Center
Schools of Medicine
United States
Zip Code
LeNoue-Newton, Michele L; Wadzinski, Brian E; Spiller, Benjamin W (2016) The three Type 2A protein phosphatases, PP2Ac, PP4c and PP6c, are differentially regulated by Alpha4. Biochem Biophys Res Commun 475:64-9
Williams, Byron C; Filter, Joshua J; Blake-Hodek, Kristina A et al. (2014) Greatwall-phosphorylated Endosulfine is both an inhibitor and a substrate of PP2A-B55 heterotrimers. Elife 3:e01695
Kamoun, Malek; Filali, Mohammed; Murray, Michael V et al. (2013) Protein phosphatase 2A family members (PP2A and PP6) associate with U1 snRNP and the spliceosome during pre-mRNA splicing. Biochem Biophys Res Commun 440:306-11
Watkins, Guy R; Wang, Ning; Mazalouskas, Matthew D et al. (2012) Monoubiquitination promotes calpain cleavage of the protein phosphatase 2A (PP2A) regulatory subunit ?4, altering PP2A stability and microtubule-associated protein phosphorylation. J Biol Chem 287:24207-15
Wang, Ning; Leung, Hung-Tat; Mazalouskas, Matthew D et al. (2012) Essential roles of the Tap42-regulated protein phosphatase 2A (PP2A) family in wing imaginal disc development of Drosophila melanogaster. PLoS One 7:e38569
LeNoue-Newton, Michele; Watkins, Guy R; Zou, Ping et al. (2011) The E3 ubiquitin ligase- and protein phosphatase 2A (PP2A)-binding domains of the Alpha4 protein are both required for Alpha4 to inhibit PP2A degradation. J Biol Chem 286:17665-71
Panosian, Timothy D; Nannemann, David P; Watkins, Guy R et al. (2011) Bacillus cereus phosphopentomutase is an alkaline phosphatase family member that exhibits an altered entry point into the catalytic cycle. J Biol Chem 286:8043-54
Widau, Ryan C; Jin, Yijun; Dixon, Shelley A et al. (2010) Protein phosphatase 2A (PP2A) holoenzymes regulate death-associated protein kinase (DAPK) in ceramide-induced anoikis. J Biol Chem 285:13827-38
McConnell, Jamie L; Watkins, Guy R; Soss, Sarah E et al. (2010) Alpha4 is a ubiquitin-binding protein that regulates protein serine/threonine phosphatase 2A ubiquitination. Biochemistry 49:1713-8
Reece, Kelie M; Mazalouskas, Matthew D; Wadzinski, Brian E (2009) The Balpha and Bdelta regulatory subunits of PP2A are necessary for assembly of the CaMKIV.PP2A signaling complex. Biochem Biophys Res Commun 386:582-7

Showing the most recent 10 out of 16 publications