Phosducin interacts with the G-protein βγ-dimer of ciliate protozoan<i>Blepharisma japonicum</i>upon illumination

Sobierajska, Katarzyna; Fabczak, Hanna; Fabczak, S.

doi:10.1242/jeb.005132

Cited by 6 publications

(8 citation statements)

References 64 publications

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“…It belongs to a eukaryotic chaperonin complex of approximately 900 kDa termed 'TCP1 ring complex' (TRiC) or chaperonin containing TCP1 (CCT) complex where TCP1 has been shown to consist of eight or nine structurally related subunits of 52-65 kDa [49][50][51][52]. Its multiple functions are to assist in the folding of some proteins including a/b-tubulin heterodimers [53] or actin [54], as well as to interact with phosducin (Pdc)-like protein (PhLP) [55] involved in the regulation of cell signalling through its interaction with the G protein bg subunit [56][57][58] and finally, with TRN1 although this interaction has not been further characterized [57].…”

Section: Signal Transduction-based Proteomicsmentioning

confidence: 99%

Chemokine receptor CCR2 undergoes transportin1‐dependent nuclear translocation

Favre¹,

Camps

Arod

et al. 2008

Proteomics

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Chemokines (CCs) are small chemoattractant cytokines involved in a wide variety of biological and pathological processes. Released by cells in the milieu, and extracellular matrix and activating signalling cascades upon binding to specific G protein-coupled receptors (GPCRs), they trigger many cellular events. In various pathologies, CCs are directly responsible for excessive recruitment of leukocytes to inflammatory sites and recent studies using chemokine receptor (CCR) antagonists permitted these molecules to reach the market for medical use. While interaction of CCs with their receptors has been extensively documented, downstream GPCR signalling cascades triggered by CC are less well understood. Given the pivotal role of chemokine receptor 2 (CCR2) in monocyte recruitment, activation and differentiation and its implication in several autoimmune-inflammatory pathologies, we searched for potential new CCR2-interacting proteins by engineering a modified CCR2 that we used as bait. Herein, we show the direct interaction of CCR2 with transportin1 (TRN1), which we demonstrate is followed by CCR2 receptor internalization. Further characterization of this novel interaction revealed that TRN1-binding to CCR2 increased upon time in agonist treated cells and promotes its nuclear translocation in a TRN1-dependent manner. Finally, we provide evidence that following translocation, the receptor localizes at the outer edge of the nuclear envelope where it is finally released from TRN1.

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Section: Signal Transduction-based Proteomicsmentioning

confidence: 99%

Chemokine receptor CCR2 undergoes transportin1‐dependent nuclear translocation

Favre¹,

Camps

Arod

et al. 2008

Proteomics

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“…5 A, B, H) while under the same light conditions, Gb formed a thin layer at the plasma membrane. 16 Therefore, in ciliates not exposed to light, Pdc and Gb did not colocalize. After a short period of light stimulation (30 s), Pdc preferentially occupied the submembrane region (Fig.…”

Section: Resultsmentioning

confidence: 91%

“…Light-induced dephosphorylation of Pdc in Blepharisma is associated with the formation of a Pdc and Gbg complex that translocates into the cell cytoplasm. 16 Therefore, we decided to determine if a decrease in Pdc phosphorylation levels due to H-89 inhibition of PKA activity in dark-adapted ciliates (Fig. 4) would produce molecular events similar to those caused by light.…”

Section: Resultsmentioning

confidence: 99%

“…These results indicate that Pdc dephosphorylation and its translocation within the cell, along with the formation of the Pdc-Gbg complex or translocated Gbg alone, may regulate the lightdependent elongation process in Blepharisma. 16 The interaction between Gbg and the potential substrate b-tubulin was examined to assess if the light-alterated Pdc phosphorylation initiate cell cytoskeleton remodelling, what may lead to changes in body shape in Blepharisma. To evaluate this hypothesis, immunocytochemical and FRET-AP analyses were performed for Gbg and b-tubulin in ciliates either adapted to darkness or exposed to illumination for different time periods.…”

Section: Resultsmentioning

confidence: 99%

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Visualization of the interaction between Gβγ and tubulin during light-induced cell elongation of Blepharisma japonicum

Sobierajska

Głos

Dąborowska

et al. 2010

Photochem Photobiol Sci

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Blepharisma japonicum ciliates display reversible cell elongation in response to lasting bright illumination. This light-induced phenomenon has been ascribed to the active sliding of the cortical microtubules of the ciliate. The detailed intracellular signaling pathway that activates the microtubule network in response to light, resulting in cell elongation, is unknown. We have previously reported that light stimulation initiates sequential molecular events consisting of a decrease in the phosphorylation of ciliate Pdc, followed by increased binding of Pdc to membrane-localised Gbetagamma and the subsequent translocation of the Pdc-Gbetagamma complex to the cytoplasm. In this study, we used selected agents known to influence protein phosphorylation to test whether alterations in Pdc phosphorylation levels by light affect ciliate shape. Behavioural analysis indicated that cell treatment with okadaic acid, an inhibitor of protein phosphatase activity, heavily abolished the effect of light on cell elongation, whereas the presence of H-89, a specific inhibitor of cAMP-dependent protein kinase (PKA) activity, had no appreciable effect on the cell length. Phosphorylation assays showed that cell incubation with H-89 mimicked light by promoting Pdc dephosphorylation and its colocalization with Gbetagamma. However, as demonstrated by FRET-AP, Pdc-Gbetagamma complex formation and changes in the length of the cell did not occur under the same conditions. Moreover, fluorescence microscopy showed localization of Gbetagamma and beta-tubulin in the same cell compartment and demonstrated that a direct interaction between these proteins occurs in cells adapted to darkness or exposed to prolonged illumination (> or = 10 min). In contrast, an opposite effect, i.e. a transient decrease in the interaction between Gbetagamma and beta-tubulin and distinct Pdc dephosphorylation, was observed in cells illuminated for short time. Under these conditions, Pdc preferentially occupies the cell submembrane region and interacts with Gbetagamma. In cells illuminated for a longer time (> or = 10 min) and despite the constant light intensity, Pdc was progressively rephosphorylated and then dissociated from Gbetagamma, relocalizing within the cell cytoplasm. The results obtained in this study suggest that alterations in Pdc phosphorylation may be involved in light-induced elongation of the Blepharisma cell body, which affects the interaction of Gbetagamma with beta-tubulin and cell cytoskeleton remodelling.

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“…Sequences were aligned using ClustalW alignment tool available from EMBL-EBI (http://www.ebi.ac. uk/Tools/clustaw/index.html) [143]. Colour code marks degree of homology between species illumination [34,38].…”

Section: Localisationmentioning

confidence: 99%

The physiological roles of phosducin: from retinal function to stress-dependent hypertension

Beetz

Hein

2010

Cell. Mol. Life Sci.

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In the time since its discovery, phosducin's functions have been intensively studied both in vivo and in vitro. Phosducin's most important biochemical feature in in vitro studies is its binding to heterotrimeric G protein βγ-subunits. Data on phosducin's in vivo relevance, however, have only recently been published but expand the range of biological actions, as shown both in animal models as well as in human studies. This review gives an overview of different aspects of phosducin biology ranging from structure, phylogeny of phosducin family members, posttranscriptional modification, biochemical features, localization and levels of expression to its physiological functions. Special emphasis will be placed on phosducin's function in the regulation of blood pressure. In the second part of this article, findings concerning cardiovascular regulation and their clinical relevance will be discussed on the basis of recently published data from gene-targeted mouse models and human genetic studies.

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Phosducin interacts with the G-protein βγ-dimer of ciliate protozoanBlepharisma japonicumupon illumination

Cited by 6 publications

References 64 publications

Chemokine receptor CCR2 undergoes transportin1‐dependent nuclear translocation

Chemokine receptor CCR2 undergoes transportin1‐dependent nuclear translocation

Visualization of the interaction between Gβγ and tubulin during light-induced cell elongation of Blepharisma japonicum

The physiological roles of phosducin: from retinal function to stress-dependent hypertension

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