Characterization of a cDNA encoding the 55 kDa B regulatory subunit of Arabidopsis protein phosphatase 2A

Abstract

Type 2A serine/threonine protein phosphatases (PP2A) are key components in the regulation of signal transduction and control of cell metabolism. The activity of these protein phosphatases is modulated by regulatory subunits. While PP2A activity has been characterized in plants, little is known about its regulation. We used the polymerase chain reaction to amplify a segment of a cDNA encoding the B regulatory subunit of PP2A from Arabidopsis. The amplified DNA fragment of 372 nucleotides was used as a probe to screen an Arabidopsis cDNA library and a full-length clone (AtB alpha) of 2.1 kbp was isolated. The predicted protein encoded by AtB alpha is 43 to 46% identical and 53 to 56% similar to its yeast and mammalian counterparts, and contains three unique regions of amino acid insertions not present in the animal B regulatory subunit. Genomic Southern blots indicate the Arabidopsis genome contains at least two genes encoding the B regulatory subunit. In addition, other plant species also contain DNA sequences homologous to the B regulatory subunit, indicating that regulation of PP2A activity by the 55 kDa B regulatory subunit is probably ubiquitous in plants. Northern blots indicate the AtB alpha mRNA accumulates in all Arabidopsis tissues examined, suggesting the protein product of the AtB alpha gene performs a basic housekeeping function in plant cells.

The Arabidopsis Homolog of Yeast TAP42 and Mammalian α4 Binds to the Catalytic Subunit of Protein Phosphatase 2A and Is Induced by Chilling

“…A 550-bp EcoRI-PstI fragment from the largest Arabidopsis cDNA positive identified in the yeast two-hybrid screen was used as a probe to screen 150,000 clones of a sizefractionated (1-2 kb) Arabidopsis cDNA library (Kieber et al, 1993), as described by Rundle et al (1995). In addition, two ESTs (82E11T7 and 143E19T7) matching the sequence of the yeast two-hybrid positive were obtained from the Arabidopsis Biological Resource Center (Ohio State University, Columbus).…”

“…Southern-blot analysis was performed as indicated in Rundle et al (1995). Blots were probed with a radiolabeled fragment spanning nucleotides 111 to 558 of the TAP46 cDNA.…”

“…RNA was separated by formaldehyde agarose electrophoresis and transferred to nylon membranes. Filters were probed as described by Rundle et al (1995). Relative RNA levels were quantified by densitometry.…”

Molecular Characterization and Evolution of the Protein Phosphatase 2A B′ Regulatory Subunit Family in Plants

“…The A regulatory subunit has a molecular mass of 65 kD, and consists of 15 imperfect repeats of 38 to 43 amino acids, through which it interacts with the PP2Ac catalytic subunit and the B regulatory subunit (Groves et al, 1999). Type B regulatory subunits of PP2A are very diverse, and can be clustered into at least three distinct groups including the 55-kD B, the 52-to 74-kD BЈ, and the 72-to 130-kD BЈЈ subunit families (Rundle et al, 1995; Corum et al, 1996; Csortos et al, 1996; McCright et al, 1996a). Each family is composed of several members with the exception of the BЈЈ subunit family (Hendrix et al, 1993a).…”

“…In Arabidopsis, the catalytic subunit of PP2 (PP2Ac) is encoded by at least five genes, each of which appears to be expressed in all tissues albeit at different levels (Ariñ o et al, 1993;Casamayor et al, 1994; Pérez-Callejó n et al, 1998). Re-garding the regulatory subunits, three genes encoding the A 65-kD subunit (Slabas et al, 1994), two genes encoding the B subunit (Rundle et al, 1995;Corum et al, 1996), and one gene encoding the BЈЈ regulatory subunit (Sato et al, 1997) have been identified. In the last years, four isoforms of the BЈ regulatory subunit of PP2A have been described in Arabidopsis, named AtBЈ␣, AtBЈ␤, AtBЈ␥ (Latorre et al, 1997), and AtBЈ␦ (Haynes et al, 1999).…”