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Mol. Biol. Cell 12 (1): 185-199

Copyright © 2001 by The American Society for Cell Biology.

Vol. 12, Issue 1, 185-199, January 2001

Methylation of the Protein Phosphatase 2A Catalytic Subunit Is Essential for Association of Balpha Regulatory Subunit But Not SG2NA, Striatin, or Polyomavirus Middle Tumor Antigen

Xing Xian Yu,* Xianxing Du,* Carlos S. Moreno,* Richard E. Green, Egon Ogris,dagger Dagger Qi Feng,dagger § Lisa Chou,dagger Monica J. McQuoid, and David C. Pallasdagger ||

Department of Biochemistry and Winship Cancer Center, Emory University School of Medicine, Atlanta, Georgia 30322.

Binding of different regulatory subunits and methylation of the catalytic (C) subunit carboxy-terminal leucine 309 are two important mechanisms by which protein phosphatase 2A (PP2A) can be regulated. In this study, both genetic and biochemical approaches were used to investigate regulation of regulatory subunit binding by C subunit methylation. Monoclonal antibodies selectively recognizing unmethylated C subunit were used to quantitate the methylation status of wild-type and mutant C subunits. Analysis of 13 C subunit mutants showed that both carboxy-terminal and active site residues are important for maintaining methylation in vivo. Severe impairment of methylation invariably led to a dramatic decrease in Balpha subunit binding but not of striatin, SG2NA, or polyomavirus middle tumor antigen (MT) binding. In fact, most unmethylated C subunit mutants showed enhanced binding to striatin and SG2NA. Certain carboxy-terminal mutations decreased Balpha subunit binding without greatly affecting methylation, indicating that Balpha subunit binding is not required for a high steady-state level of C subunit methylation. Demethylation of PP2A in cell lysates with recombinant PP2A methylesterase greatly decreased the amount of C subunit that could be coimmunoprecipitated via the Balpha subunit but not the amount that could be coimmunoprecipitated with Aalpha subunit or MT. When C subunit methylation levels were greatly reduced in vivo, Balpha subunits were found complexed exclusively to methylated C subunits, whereas striatin and SG2NA in the same cells bound both methylated and unmethylated C subunits. Thus, C subunit methylation is critical for assembly of PP2A heterotrimers containing Balpha subunit but not for formation of heterotrimers containing MT, striatin, or SG2NA. These findings suggest that methylation may be able to selectively regulate the association of certain regulatory subunits with the A/C heterodimer.


* These authors contributed equally to this work.

dagger Portions of this work were performed while these investigators were in the Division of Cellular and Molecular Biology, Dana-Farber Cancer Institute, and Department of Pathology, Harvard Medical School, Boston, MA.

Dagger E.O. was supported by an Erwin Schrödinger Fellowship from Austrian Fonds zur Förderung der Wissenschaftlichen Forschung. Present address: Institute of Molecular Biology, Vienna Biocenter, University of Vienna, A-1030 Vienna, Austria.

§ Present address: Oklahoma University Health Science Center, Oklahoma City, OK 73106.

|| Corresponding author. E-mail address: dpallas{at}emory.edu.


Molecular Biology of the Cell
Vol. 12, 185-199, January 2001
Copyright © 2001 by The American Society for Cell Biology


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