Leucine Carboxyl Methyltransferase 1(LCMT-1) Methylates Protein Phosphatase 4(PP4) and Protein Phosphatase 6(PP6) and Differentially Regulates the Stable Formation of Different PP4 Holoenzymes

• Type of Material: Article
• Authors: Juyeon Hwang, Emory University; Jocelyn A. Lee, Emory University; David Pallas, Emory University
• Language: English
• Date: 2016-09-30
• Publisher: American Society for Biochemistry and Molecular Biology
• Publication Version: Final Published Version
• Copyright Statement: © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.
• Final Published Version (URL): http://dx.doi.org/10.1074/jbc.M116.739920
• Title of Journal or Parent Work: Journal of Biological Chemistry
• ISSN: 0021-9258
• Volume: 291
• Issue: 40
• Start Page: 21008
• End Page: 21019
• Grant/Funding Information: Jocelyn A. Lee is supported by National Institutes of Health Predoctoral Fellowship Grant 5F31CA123640 (NCI).; This work was also supported by an award from the Emory University Research Committee.; This work was supported by National Institutes of Health Grant R01 CA57327 (NCI).
• Keywords: MIDDLE TUMOR-ANTIGEN; Life Sciences & Biomedicine; DEPHOSPHORYLATES GAMMA-H2AX; SACCHAROMYCES-CEREVISIAE; ALPHA4 PROTEIN; BOVINE BRAIN; Lcmt-1; PP6; Protein phosphorylation; Methyltransferase; 2A CATALYTIC SUBUNIT; Biochemistry & Molecular Biology; Protein methylation; Science & Technology; PP2A; Protein complex; DNA-DAMAGE CHECKPOINT; IN-VIVO; Protein phosphatase 2 (PP2A); ASSOCIATION; Signaling
• Research Categories: Biology, Cell; Chemistry, Biochemistry

Abstract

The protein phosphatase 2A (PP2A) subfamily of phosphatases, PP2A, PP4, and PP6, are multifunctional serine/threonine protein phosphatases involved in many cellular processes. Carboxyl methylation of the PP2A catalytic subunit (PP2Ac) C-terminal leucine is regulated by the opposing activities of leucine carboxyl methyltransferase 1 (LCMT-1) and protein phosphatase methylesterase 1 (PME-1) and regulates PP2A holoenzyme formation. The site of methylation on PP2Ac is conserved in the catalytic subunits of PP4 and PP6, and PP4 is also methylated on that site, but the identities of the methyltransferase enzyme for PP4 are not known. Whether PP6 is methylated is also not known. Here we use antibodies specific for the unmethylated phosphatases to show that PP6 is carboxyl-methylated and that LCMT-1 is the major methyltransferase for PP2A, PP4, and PP6 in mouse embryonic fibroblasts (MEFs). Analysis of PP2A and PP4 complexes by blue native polyacrylamide gel electrophoresis (BN-PAGE) indicates that PP4 holoenzyme complexes, like those of PP2A, are differentially regulated by LCMT-1, with the PP4 regulatory subunit 1 (PP4R1)-containing PP4 complex being the most dramatically affected by the LCMT-1 loss. MEFs derived from LCMT-1 knock-out mouse embryos have reduced levels of PP2A B regulatory subunit and PP4R1 relative to control MEFs, indicating that LCMT-1 is important for maintaining normal levels of these subunits. Finally, LCMT-1 homozygous knock-out MEFs exhibited hyperphosphorylation of HDAC3, a reported target of the methylation-dependent PP4R1-PP4c complex. Collectively, our data suggest that LCMT-1 coordinately regulates the carboxyl methylation of PP2A-related phosphatases and, consequently, their holoenzyme assembly and function.