GENOMICS Vol. 79, Number 3, March 2002 Copyright © 2002 Elsevier Science (USA). All rights reserved. 0888-7543/02 $35.00 344 Article doi:10.1006/geno.2002.6721, available online at http://www.idealibrary.com on IDEAL INTRODUCTION Many cellular processes are dependent on the phosphoryla- tion state of key regulatory proteins. The serine/threonine phosphatase, protein phosphatase 2A (PP2A), affects the phosphorylation status of many proteins and has been linked to the regulation of signal transduction pathways, neoplastic transformation, cell cycle progression, DNA replication/tran- scription, and embryonic development [1,2]. The core enzyme of PP2A is a heterodimer consisting of a catalytic (C) and a structural (A) subunit. Differential association of regulatory B subunits with the AC core dimer confers enzymatic speci- ficity and intracellular targeting of the heterotrimeric PP2A holoenzyme [2,3]. There are three distinct families of cellular regulatory subunits (B, B', and B”) with multiple isoforms within each [4]. The B' subunits, also designated as B56, are the most diverse, consisting of  (PPP2R5A),  (PPP2R5B),  (PPP2R5C),  (PPP2R5D), and  (PPP2R5E) isoforms and three differentially spliced PPP2R5C variants (1, 2, and 3) [5,6]. The human B56 genes were mapped by cytogenetic analy- sis to chromosomes 1q41 (PPP2R5A), 11q12 (PPP2R5B), 3p21 (PPP2R5C), 6p21 (PPP2R5D), and 7p11 (PPP2R5E) [7]. Subsequently, Généthon and the Whitehead Institute Genomic Organization and Mapping of the Gene Encoding the PP2A B56 Regulatory Subunit Sabeeha Muneer, 1 Vivek Ramalingam, 1 Robert Wyatt, 4 Roger A. Schultz, 4 John D. Minna, 1,2,3 and Craig Kamibayashi 1,2,* 1 Hamon Center for Therapeutic Oncology Research, Departments of 2 Pharmacology, 3 Internal Medicine, and 4 Pathology, University of Texas Southwestern Medical Center, 6000 Harry Hines Blvd., Dallas, Texas 75390, USA * To whom correspondence and reprint requests should be addressed. Fax: (214) 648-4940. E-mail: Craig.Kamibayashi@UTSouthwestern.edu. Protein phosphatase 2A (PP2A) is a major serine/threonine phosphatase that regulates a wide variety of cellular processes. The enzymatic activity and intracellular localization of PP2A are determined by three distinct families of cellular regulatory subunits (B, B', and B”). The B' subunit, also known as B56, is the most diverse, consisting of five isoforms (, , , , and ). The gene encoding B56 has been designated as PPP2R5C and encodes three differentially spliced variants: B561, -2, and -3. However, conflicting chromosomal loci have been reported in human genomic databases. The original cytogenetic mapping placed the gene on chromosome 3p21.3, whereas subsequent studies using radiation hybrid analysis localized PPP2R5C to chromosome 14q. In this study, by radiation hybrid mapping, FISH analysis, BAC clone sequencing, and RT-PCR analysis, we show that the functional gene PPP2R5C exists at 14q32.2 and gives rise to three splicing variants, B561, -2, and -3, whereas a nonfunc- tional B561 pseudogene, PPP2R5CP, is present at 3p21.3. We also report the genomic organ- ization of both the functional gene and the pseudogene. Key Words: PP2A, genomic organization, RH mapping confirmed these loci by radiation hybrid (RH) analyses which were deposited into the UniGene genomic database. However, the RH mapping placed PPP2R5C on both 3p21 and the telomere of 14q. The original description that PPP2R5C was differentially spliced was based on the presence of a conserved splice site in the 3’ end of the coding region of the mouse B561 and human B561, -2, and -3 cDNAs [8]. The untranslated regions (UTRs) of B562 and -3 were iden- tical, indicating that they were products of the same gene; however, it was not possible to make the same conclusion for B561. Although the 5' UTR of B561 was identical to that of -2 and -3, the 3' UTR differed, raising the possibility that a gene duplication event had occurred giving rise to two sep- arate genes. In evaluating the role of the PP2A genes as tumor sup- pressor genes, we have observed frequent 3p allele loss and developed other data suggesting that the B56 subunits can suppress tumor cell growth (unpublished data). Further, when our mutational analysis of the B56 subunits in human tumors showed alterations (unpublished data), we were unable to differentiate whether the mutations occurred at the 14q or 3p sites, requiring us to determine the genomic structure.