High Complexity in the Expression of the B′ Subunit of Protein Phosphatase 2A

doi:10.1074/jbc.271.5.2578

High Complexity in the Expression of the B′ Subunit of Protein Phosphatase 2A

EVIDENCE FOR THE EXISTENCE OF AT LEAST SEVEN NOVEL ISOFORMS (*)

From the Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana 46202-5122

**To whom correspondence should be addressed:
Dept. of Biochemistry and Molecular Biology, Indiana University School of Medicine, 635 Barnhill Dr., Indianapolis, IN 46202-5122
. Tel.: 317-274-1585; Fax: 317-274-4686.

↵^§ Present address: Dept. of Medical Chemistry, University School of Medicine, Debrecen, Bem ter 18/B, Debrecen, Hungary, 4026.
↵^¶ Present address: Dept. of Biochemistry, Faculty of Biotechnology, Medical University of Gdansk, ul. Debinki 1, 80-210 Gdansk, Poland.

Abstract

Association of the catalytic subunit (C2) with a variety of regulatory subunits is believed to modulate the activity and specificity of protein phosphatase 2A (PP2A). In this study we report the cloning and expression of a new family of B-subunit, the B′, associated with the PP2A form. Polymerase chain reactions and cDNA library screening have identified at least seven cDNA isotypes, designated α, β1, β2, β3, β4, , and . The different β subtypes appear to be generated by alternative splicing. The deduced amino acid sequences of the α, β2, β3, β4 and isoforms predict molecular weights of 57,600, 56,500, 60,900, 52,500, and 68,000, respectively. The proteins are 60-80% identical and differ mostly at their termini. Two of the isoforms, B′β3 and B′, contain a bipartite nuclear localization signal in their COOH terminus. No homology was found with other B- or Brelated subunits. Northern analyses indicate a tissuespecific expression of the isoforms. Expression of B′α protein in Escherichia coli generated a polypeptide of 53 kDa, similar to the size of the B′ subunit present in the purified PP2A. The recombinant protein was recognized by antibody raised against native B′ and interacted with the dimeric PP2A (A•C2) to generate a trimeric phosphatase. The deduced amino acid sequences of the B′ isoforms show significant homology to mammalian, fungal, and plant nucleotide sequences of unknown function present in the data bases. Notably, a high degree of homology (55-66%) was found with a yeast gene, RTS1, encoding a multicopy suppressor of a rox3 mutant. Our data indicate that at least seven B′ subunit isoforms may participate in the generation of a large number of PP2A holoenzymes that may be spatially and/or functionally targeted to different cellular processes.

Footnotes

↵* This work was supported by National Institutes of Health Grant P01 HL6308, Project 4. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank(™)/EMBL Data Bank with accession number(s) U37769 (B′α), U37770 (B′β1), U38190 (B′β2), U38191 (B′β3), U38192 (B′β4), U38193 and U38195 (B′), and U38194 (B′).
↵¹ The abbreviations used are:

PP2A

protein phosphatase type 2A

PAGE

polyacrylamide gel electrophoresis

PCR

polymerase chain reaction

bp

base pair(s)

NTA

nitrilotriacetic acid

kb

kilobase(s).
↵²M. Takeda, personal communication.
↵³C. C. Evangelista, A. M. Rodriguez Torres, M. P. Limbach, and R. S. Zitomer, unpublished results.
↵⁴Y. Zhao, G. Boguslawski, R. Zitomer, and A. A. DePaoli-Roach, unpublished results.
- Received September 29, 1995.
- Revision received November 28, 1995.