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Protein phosphatase-1α interacts with and dephosphorylates polycystin-1.

Parnell SC, Puri S, Wallace DP, Calvet JP - PLoS ONE (2012)

Bottom Line: We have now determined that polycystin-1 is a regulatory subunit for protein phosphatase-1α (PP1α).To determine whether PP1α dephosphorylates polycystin-1, a PKA-phosphorylated GST-polycystin-1 fusion protein was shown to be dephosphorylated by PP1α but not by PP2B (calcineurin).The results suggest that polycystin-1 forms a holoenzyme complex with PP1α via a conserved PP1-binding motif within the polycystin-1 C-tail, and that PKA-phosphorylated polycystin-1 serves as a substrate for the holoenzyme.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America. sparnell@kumc.edu

ABSTRACT
Polycystin signaling is likely to be regulated by phosphorylation. While a number of potential protein kinases and their target phosphorylation sites on polycystin-1 have been identified, the corresponding phosphatases have not been extensively studied. We have now determined that polycystin-1 is a regulatory subunit for protein phosphatase-1α (PP1α). Sequence analysis has revealed the presence of a highly conserved PP1-interaction motif in the cytosolic, C-terminal tail of polycystin-1; and we have shown that transfected PP1α specifically co-immunoprecipitates with a polycystin-1 C-tail construct. To determine whether PP1α dephosphorylates polycystin-1, a PKA-phosphorylated GST-polycystin-1 fusion protein was shown to be dephosphorylated by PP1α but not by PP2B (calcineurin). Mutations within the PP1-binding motif of polycystin-1, including an autosomal dominant polycystic kidney disease (ADPKD)-associated mutation, significantly reduced PP1α-mediated dephosphorylation of polycystin-1. The results suggest that polycystin-1 forms a holoenzyme complex with PP1α via a conserved PP1-binding motif within the polycystin-1 C-tail, and that PKA-phosphorylated polycystin-1 serves as a substrate for the holoenzyme.

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Related in: MedlinePlus

Model of PC1-PP1α holoenzyme function.In this model we speculate that PP1 regulates the phosphorylation status of PC1 and PC1-interacting proteins, and that disruption of the PC1-PP1 holoenzyme complex may lead to altered signaling and cystogenesis due to misregulation of protein phosphorylation.
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pone-0036798-g007: Model of PC1-PP1α holoenzyme function.In this model we speculate that PP1 regulates the phosphorylation status of PC1 and PC1-interacting proteins, and that disruption of the PC1-PP1 holoenzyme complex may lead to altered signaling and cystogenesis due to misregulation of protein phosphorylation.

Mentions: In Figure 7, we present a model of PC1-PP1 holoenzyme function whereby signaling events downstream of PC1 are regulated by the competing actions of PP1 and cellular kinases acting on PC1 itself or other, putative substrates of the holoenzyme. For example, PP1 may serve as a molecular switch that antagonizes PC1-mediated signaling initiated by events such as PKA-mediated phosphorylation of PC1. Alternatively, PP1 may regulate the phosphorylation of PC1-interacting proteins such as PC2, a protein whose localization and activation are known to be affected by phosphorylation [45], [46], [47], [48]. In our model, we predict that perturbation of PC1-PP1 holoenzyme function, either by complete loss of PC1 or by PC1 mutations that affect the activity of PP1 in complex with PC1 (e.g. RVxF mutations, Figure 6), would lead to disregulated phosphorylation and cell signaling that could potentially initiate or exacerbate cystogenesis. In summary, we have identified PC1 both as a regulatory subunit and as a substrate for PP1α. We propose that PC1 and PP1α form a holoenzyme complex in which the two proteins stably interact and are poised to dephosphorylate substrates required for normal cellular functions. The identification of PC1 mutations that interfere with the activity of PP1α in complex with PC1 should facilitate efforts to understand the role of PP1α in PC1-mediated signaling as well as the role of PKA phosphorylation in the function of PC1.


Protein phosphatase-1α interacts with and dephosphorylates polycystin-1.

Parnell SC, Puri S, Wallace DP, Calvet JP - PLoS ONE (2012)

Model of PC1-PP1α holoenzyme function.In this model we speculate that PP1 regulates the phosphorylation status of PC1 and PC1-interacting proteins, and that disruption of the PC1-PP1 holoenzyme complex may lead to altered signaling and cystogenesis due to misregulation of protein phosphorylation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3366979&req=5

pone-0036798-g007: Model of PC1-PP1α holoenzyme function.In this model we speculate that PP1 regulates the phosphorylation status of PC1 and PC1-interacting proteins, and that disruption of the PC1-PP1 holoenzyme complex may lead to altered signaling and cystogenesis due to misregulation of protein phosphorylation.
Mentions: In Figure 7, we present a model of PC1-PP1 holoenzyme function whereby signaling events downstream of PC1 are regulated by the competing actions of PP1 and cellular kinases acting on PC1 itself or other, putative substrates of the holoenzyme. For example, PP1 may serve as a molecular switch that antagonizes PC1-mediated signaling initiated by events such as PKA-mediated phosphorylation of PC1. Alternatively, PP1 may regulate the phosphorylation of PC1-interacting proteins such as PC2, a protein whose localization and activation are known to be affected by phosphorylation [45], [46], [47], [48]. In our model, we predict that perturbation of PC1-PP1 holoenzyme function, either by complete loss of PC1 or by PC1 mutations that affect the activity of PP1 in complex with PC1 (e.g. RVxF mutations, Figure 6), would lead to disregulated phosphorylation and cell signaling that could potentially initiate or exacerbate cystogenesis. In summary, we have identified PC1 both as a regulatory subunit and as a substrate for PP1α. We propose that PC1 and PP1α form a holoenzyme complex in which the two proteins stably interact and are poised to dephosphorylate substrates required for normal cellular functions. The identification of PC1 mutations that interfere with the activity of PP1α in complex with PC1 should facilitate efforts to understand the role of PP1α in PC1-mediated signaling as well as the role of PKA phosphorylation in the function of PC1.

Bottom Line: We have now determined that polycystin-1 is a regulatory subunit for protein phosphatase-1α (PP1α).To determine whether PP1α dephosphorylates polycystin-1, a PKA-phosphorylated GST-polycystin-1 fusion protein was shown to be dephosphorylated by PP1α but not by PP2B (calcineurin).The results suggest that polycystin-1 forms a holoenzyme complex with PP1α via a conserved PP1-binding motif within the polycystin-1 C-tail, and that PKA-phosphorylated polycystin-1 serves as a substrate for the holoenzyme.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America. sparnell@kumc.edu

ABSTRACT
Polycystin signaling is likely to be regulated by phosphorylation. While a number of potential protein kinases and their target phosphorylation sites on polycystin-1 have been identified, the corresponding phosphatases have not been extensively studied. We have now determined that polycystin-1 is a regulatory subunit for protein phosphatase-1α (PP1α). Sequence analysis has revealed the presence of a highly conserved PP1-interaction motif in the cytosolic, C-terminal tail of polycystin-1; and we have shown that transfected PP1α specifically co-immunoprecipitates with a polycystin-1 C-tail construct. To determine whether PP1α dephosphorylates polycystin-1, a PKA-phosphorylated GST-polycystin-1 fusion protein was shown to be dephosphorylated by PP1α but not by PP2B (calcineurin). Mutations within the PP1-binding motif of polycystin-1, including an autosomal dominant polycystic kidney disease (ADPKD)-associated mutation, significantly reduced PP1α-mediated dephosphorylation of polycystin-1. The results suggest that polycystin-1 forms a holoenzyme complex with PP1α via a conserved PP1-binding motif within the polycystin-1 C-tail, and that PKA-phosphorylated polycystin-1 serves as a substrate for the holoenzyme.

Show MeSH
Related in: MedlinePlus