Structure and transcriptional regulation of protein phosphatase 2A catalytic subunit genes [Erratum to document cited in CA114(3):18572v]

Abstract Mutations in the gene coding for the multi-domain protein leucine-rich repeat kinase 2 (LRRK2) are the leading cause of genetically inherited Parkinson’s disease (PD). Two of the common found mutations are the R1441C and G2019S. In this study we identified protein phosphatase 2A (PP2A) as an interacting partner of LRRK2. We were able to demonstrate that the Ras of complex protein (ROC) domain is sufficient to interact with the three subunits of PP2A in human neuroblastoma SH-SY5Y cells and in HeLa cells. The alpha subunit of PP2A is interacting with LRRK2 in the perinuclear region of HeLa cells. Silencing the catalytic subunit of PP2A by shRNA aggravated cellular degeneration induced by the pathogenic R1441C-LRRK2 mutant expressed in neuroblastoma SH-SY5Y cells. A similar enhancement of apoptotic nuclei was observed by downregulation of the catalytic subunit of PP2A in cultured cortical cells derived from neurons overexpressing the pathogenic mutant G2019S-LRRK2. Conversely, pharmacological activation of PP2A by sodium selenate showed a partial neuroprotection from R1441C-LRRK2-induced cellular degeneration. All these data suggest that PP2A is a new interacting partner of LRRK2 and reveal the importance of PP2A as a potential therapeutic target in PD.

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A Novel Interaction of the Catalytic Subunit of Protein Phosphatase 2A with the Adaptor Protein CIN85 Suppresses Phosphatase Activity and Facilitates Platelet Outside-in αIIbβ3 Integrin Signaling

Journal of Biological Chemistry ◽

10.1074/jbc.m115.704296 ◽

2016 ◽

Vol 291 (33) ◽

pp. 17360-17368 ◽

Cited By ~ 2

Author(s):

Tanvir Khatlani ◽

Subhashree Pradhan ◽

Qi Da ◽

Tanner Shaw ◽

Vladimir L. Buchman ◽

...

Keyword(s):

Phosphatase Activity ◽

Protein Interactions ◽

Protein Phosphatase ◽

Thrombus Formation ◽

Protein Phosphatase 2A ◽

Adaptor Protein ◽

Fibrin Clot ◽

Catalytic Subunit ◽

Clot Retraction ◽

Phosphatase 2A

The transduction of signals generated by protein kinases and phosphatases are critical for the ability of integrin αIIbβ3 to support stable platelet adhesion and thrombus formation. Unlike kinases, it remains unclear how serine/threonine phosphatases engage the signaling networks that are initiated following integrin ligation. Because protein-protein interactions form the backbone of signal transduction, we searched for proteins that interact with the catalytic subunit of protein phosphatase 2A (PP2Ac). In a yeast two-hybrid study, we identified a novel interaction between PP2Ac and an adaptor protein CIN85 (Cbl-interacting protein of 85 kDa). Truncation and alanine mutagenesis studies revealed that PP2Ac binds to the P3 block (396PAIPPKKPRP405) of the proline-rich region in CIN85. The interaction of purified PP2Ac with CIN85 suppressed phosphatase activity. Human embryonal kidney 293 αIIbβ3 cells overexpressing a CIN85 P3 mutant, which cannot support PP2Ac binding, displayed decreased adhesion to immobilized fibrinogen. Platelets contain the ∼85 kDa CIN85 protein along with the PP2Ac-CIN85 complex. A myristylated cell-permeable peptide derived from residues 395–407 of CIN85 protein (P3 peptide) disrupted the platelet PP2Ac-CIN85 complex and decreased αIIbβ3 signaling dependent functions such as platelet spreading on fibrinogen and thrombin-mediated fibrin clot retraction. In a phospho-profiling study P3 peptide treated platelets also displayed decreased phosphorylation of several signaling proteins including Src and GSK3β. Taken together, these data support a role for the novel PP2Ac-CIN85 complex in supporting integrin-dependent platelet function by dampening the phosphatase activity.

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Regulation by glucose and calcium of the carboxylmethylation of the catalytic subunit of protein phosphatase 2A in insulin-secreting INS-1 cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00587.2003 ◽

2004 ◽

Vol 286 (6) ◽

pp. E1032-E1041 ◽

Cited By ~ 24

Author(s):

Rengasamy Palanivel ◽

Rajakrishnan Veluthakal ◽

Anjaneyulu Kowluru

Keyword(s):

Insulin Secretion ◽

Glucose Metabolism ◽

Protein Phosphatase ◽

Protein Phosphatase 2A ◽

Catalytic Subunit ◽

Inhibitory Effects ◽

Β Cells ◽

Intact Cells ◽

Phosphatase 2A ◽

Glucose Metabolites

Previously, we reported that the catalytic subunit of protein phosphatase 2A (PP2Ac) undergoes carboxylmethylation (CML) at its COOH-terminal leucine, and that inhibitors of such a posttranslational modification markedly attenuate nutrient-induced insulin secretion from isolated β-cells. More recent studies have suggested direct inhibitory effects of glucose metabolites on PP2A activity in isolated β-cells, implying that inhibition of PP2A leads to stimulation of insulin secretion. Because the CML of PP2Ac has been shown to facilitate the holoenzyme assembly and subsequent functional activation of PP2A, we investigated putative regulation by glucose of the CML of PP2Ac in insulin-secreting (INS)-1 cells. Our data indicated a marked inhibition by specific intermediates of glucose metabolism (e.g., citrate and phospho enolpyruvate) of the CML of PP2Ac in INS-1 cell lysates. Such inhibitory effects were also demonstrable in intact cells by glucose. Mannoheptulose, an inhibitor of glucose metabolism, completely prevented inhibitory effects of glucose on the CML of PP2Ac. Moreover, glucose-mediated inhibition of the CML of PP2Ac was resistant to diazoxide, suggesting that glucose metabolism and the generation of glucose metabolites might control inhibition of the CML of PP2Ac. A membrane-depolarizing concentration of KCl also induced inhibition of the CML of PP2Ac in intact INS cells. On the basis of these data, we propose that glucose metabolism and increase in intracellular calcium facilitate inhibition of the CML of PP2Ac, resulting in functional inactivation of PP2A. This, in turn, might retain the key signaling proteins of the insulin exocytotic cascade in their phosphorylated state, leading to stimulated insulin secretion.

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Structural basis for PTPA interaction with the invariant C-terminal tail of PP2A

Biological Chemistry ◽

10.1515/hsz-2014-0106 ◽

2014 ◽

Vol 395 (7-8) ◽

pp. 881-889 ◽

Cited By ~ 12

Author(s):

Christian Löw ◽

Esben M. Quistgaard ◽

Michael Kovermann ◽

Madhanagopal Anandapadamanaban ◽

Jochen Balbach ◽

...

Keyword(s):

Signaling Pathways ◽

Posttranslational Modifications ◽

Protein Phosphatase ◽

Protein Phosphatase 2A ◽

Catalytic Subunit ◽

Structural Basis ◽

Interaction Site ◽

Interaction Partners ◽

Phosphatase 2A ◽

Additional Interaction

Abstract Protein phosphatase 2A (PP2A) is a highly abundant heterotrimeric Ser/Thr phosphatase involved in the regulation of a variety of signaling pathways. The PP2A phosphatase activator (PTPA) is an ATP-dependent activation chaperone, which plays a key role in the biogenesis of active PP2A. The C-terminal tail of the catalytic subunit of PP2A is highly conserved and can undergo a number of posttranslational modifications that serve to regulate the function of PP2A. Here we have studied structurally the interaction of PTPA with the conserved C-terminal tail of the catalytic subunit carrying different posttranslational modifications. We have identified an additional interaction site for the invariant C-terminal tail of the catalytic subunit on PTPA, which can be modulated via posttranslational modifications. We show that phosphorylation of Tyr307PP2A-C or carboxymethylation of Leu309PP2A-C abrogates or diminishes binding of the C-terminal tail, whereas phosphorylation of Thr304PP2A-C is of no consequence. We suggest that the invariant C-terminal residues of the catalytic subunit can act as affinity enhancer for different PP2A interaction partners, including PTPA, and a different ‘code’ of posttranslational modifications can favour interactions to one subunit over others.

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