Functional expression of human and Arabidopsis protein phosphatase 2A in Saccharomyces cerevisiae and isolation of dominant-defective mutants

Partial purification and biochemical characterization of a heteromeric protein phosphatase 2A holoenzyme from maize ( Zea mays L.) leaves that dephosphorylates C 4 phospho enol pyruvate carboxylase

“…The results indicate that the C 4 -leaf holoenzyme is analogous to other eukaryotic PP2As in being a 170-kDa heteromer comprised of a core PP2Ac-A heterodimer (103 kDa) complexed with a putative, 74-kDa B-type subunit. These ®ndings are consistent with the burgeoning molecular data documenting the presence of numerous C-, A-, and B-like subunit genes in plants (Smith and Walker 1996;DerueÁ re et al 1999;Haynes et al 1999;Lizotte et al 1999;Thakore et al 1999;Luan 2000), and the recent biochemical evidence for the existence of PP2A heteromers in various heterotrophic plant tissues (Awotunde et al 2000;Camoni et al 2000;To th et al 2000). Perhaps the most notable features of this C 4 -leaf PP2A holoenzyme relate to (i) its lack of strict substrate speci®city for PEPC, even though it was subjected to ®nal anity-puri®cation on a thiophosphorylated C 4 PEPC-peptide column; (ii) the absence of any striking light/dark eects on its apparent molecular mass, component core subunits and activity against C 4 PEPC, in marked contrast to the opposing activity of the dedicated PEPC-kinase in C 4 and CAM leaves (Chollet et al 1996;Vidal and Chollet 1997;Nimmo 2000); and (iii) its in vitro regulation by certain C 4 PEPC-related metabolites, including Glc6P, L-malate and PEP, but not others (i.e., glycine).…”

“…V5981). This latter rabbit polyclonal antibody was directed against the 25 Cterminal residues of human liver PP2Ac, a domain that is highly conserved in eukaryotes as distantly related as yeast, Arabidopsis, and human (Lizotte et al 1999). MC-LR and okadaic acid were obtained from either Sigma or ICN, and [c-32 P]ATP (74 MBq/ml, 111 GBq/mmol) was from Amersham Pharmacia Biotech.…”

Protein phosphatase 1 positively regulates stomatal opening in response to blue light in Vicia faba

“…The His residue in the catalytic site (highlighted in Fig. 1 A) was replaced by Asn to abolish PP1c activity (24); we expected that the mutant functioned as a dominant negative form in vivo, similarly as in the case of PP2A (20,21). We prepared recombinant VfPP1c-1 and VfPP1c-1-H137N and found that the activity was lost completely in the latter.…”

“…Mutational studies have shown that the expression of a dominant negative-type mutant of the PP2A catalytic subunit causes growth defects and reduction of PP2A activity without affecting its holoenzyme assembly in yeast (20,21). In Drosophila cells, overexpression of inhibitor-2, which inhibits PP1c activity as a regulatory subunit, specifically reduces PP1 activity without affecting that of PP2A (22).…”

Abolition of Cyclin-Dependent Kinase Inhibitor p16Ink4a and p21Cip1/Waf1 Functions Permits Ras-Induced Anchorage-Independent Growth in Telomerase-Immortalized Human Fibroblasts

“…PP2A is a heterotrimeric serine/threonine phosphatase with numerous cellular targets and biological functions (23). Although PP2A is essential for viability (13,15,33), in a general sense its activity has antiproliferative effects (23). Its targets include (among many others) the mitogen-activated protein kinase pathway (63,64), G 1 cyclin-dependent kinases (79), p70 S6 protein kinase (77), mitochondrial Bcl2 (54), and Mdm2 (42).…”