Dataset Information

Protein phosphatase 2A dephosphorylates CaBP4 and regulates CaBP4 function.

ABSTRACT: PURPOSE:CaBP4 is a neuronal Ca(2+)-binding protein that is expressed in the retina and in the cochlea, and is essential for normal photoreceptor synaptic function. CaBP4 is phosphorylated by protein kinase C zeta (PKC?) in the retina at serine 37, which affects its interaction with and modulation of voltage-gated Ca(v)1 Ca(2+) channels. In this study, we investigated the potential role and functional significance of protein phosphatase 2A (PP2A) in CaBP4 dephosphorylation. METHODS:The effect of protein phosphatase inhibitors, light, and overexpression of PP2A subunits on CaBP4 dephosphorylation was measured in in vitro assays. Pull-down experiments using retinal or transfected HEK293 cell lysates were used to investigate the association between CaBP4 and PP2A subunits. Electrophysiologic recordings of cotransfected HEK293 cells were performed to analyze the effect of CaBP4 dephosphorylation in modulating Ca(v)1.3 currents. RESULTS:PP2A inhibitors, okadaic acid (OA), and fostriecin, but not PP1 selective inhibitors, NIPP-1, and inhibitor 2, block CaBP4 dephosphorylation in retinal lysates. Increased phosphatase activity in light-dependent conditions reverses phosphorylation of CaBP4 by PKC?. In HEK293 cells, overexpression of PP2A enhances the rate of dephosphorylation of CaBP4. In addition, inhibition of protein phosphatase activity by OA increases CaBP4 phosphorylation and potentiates the modulatory effect of CaBP4 on Ca(v)1.3 Ca(2+) channels in HEK293T cells. CONCLUSIONS:This study provides evidence that CaBP4 is dephosphorylated by PP2A in the retina. Our findings reveal a novel role for protein phosphatases in regulating CaBP4 function in the retina, which may fine tune presynaptic Ca(2+) signals at the photoreceptor synapse.

SUBMITTER: Haeseleer F

PROVIDER: S-EPMC3575156 | biostudies-literature | 2013 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Protein phosphatase 2A dephosphorylates CaBP4 and regulates CaBP4 function.

Haeseleer Françoise F Sokal Izabela I Gregory Frederick D FD Lee Amy A

Investigative ophthalmology & visual science 20130201 2

<h4>Purpose</h4>CaBP4 is a neuronal Ca(2+)-binding protein that is expressed in the retina and in the cochlea, and is essential for normal photoreceptor synaptic function. CaBP4 is phosphorylated by protein kinase C zeta (PKCζ) in the retina at serine 37, which affects its interaction with and modulation of voltage-gated Ca(v)1 Ca(2+) channels. In this study, we investigated the potential role and functional significance of protein phosphatase 2A (PP2A) in CaBP4 dephosphorylation.<h4>Methods</h4 ...[more]

PMID: 23341017

Similar Datasets

Project description:BackgroundInvadopodia are actin-based cell-membrane protrusions associated with the extracellular matrix degradation accompanying cancer invasion. The elucidation of the molecular mechanisms leading to invadopodia formation and activity is central for the prevention of tumor spreading and growth. Protein tyrosine kinases such as Src are known to regulate invadopodia assembly, little is however known on the role of protein tyrosine phosphatases in this process. Among these enzymes, we have selected the tyrosine phosphatase Shp1 to investigate its potential role in invadopodia assembly, due to its involvement in cancer development.MethodsCo-immunoprecipitation and immunofluorescence studies were employed to identify novel substrate/s of Shp1AQ controlling invadopodia activity. The phosphorylation level of cortactin, the Shp1 substrate identified in this study, was assessed by immunoprecipitation, in vitro phosphatase and western blot assays. Short interference RNA and a catalytically-dead mutant of Shp1 expressed in A375MM melanoma cells were used to evaluate the role of the specific Shp1-mediated dephosphorylation of cortactin. The anti-invasive proprieties of glycerophosphoinositol, that directly binds and regulates Shp1, were investigated by extracellular matrix degradation assays and in vivo mouse model of metastasis.ResultsThe data show that Shp1 was recruited to invadopodia and promoted the dephosphorylation of cortactin at tyrosine 421, leading to an attenuated capacity of melanoma cancer cells to degrade the extracellular matrix. Controls included the use of short interference RNA and catalytically-dead mutant that prevented the dephosphorylation of cortactin and hence the decrease the extracellular matrix degradation by melanoma cells. In addition, the phosphoinositide metabolite glycerophosphoinositol facilitated the localization of Shp1 at invadopodia hence promoting cortactin dephosphorylation. This impaired invadopodia function and tumor dissemination both in vitro and in an in vivo model of melanomas.ConclusionThe main finding here reported is that cortactin is a specific substrate of the tyrosine phosphatase Shp1 and that its phosphorylation/dephosphorylation affects invadopodia formation and, as a consequence, the ability of melanoma cells to invade the extracellular matrix. Shp1 can thus be considered as a regulator of melanoma cell invasiveness and a potential target for antimetastatic drugs. Video abstract.

Dataset Information

Protein phosphatase 2A dephosphorylates CaBP4 and regulates CaBP4 function.

Publications

Protein phosphatase 2A dephosphorylates CaBP4 and regulates CaBP4 function.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets