scholarly journals Dissecting Gq/11-Mediated Plasma Membrane Translocation of Sphingosine Kinase-1

Cells ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 2201
Author(s):  
Kira Vanessa Blankenbach ◽  
Ralf Frederik Claas ◽  
Natalie Judith Aster ◽  
Anna Katharina Spohner ◽  
Sandra Trautmann ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Inositol Phosphate ◽  
Sphingosine Kinase ◽  
Binding Motif ◽  
Sphingosine Kinase 1 ◽  
Nucleotide Exchange ◽  
Ph Domain ◽  
Wild Type ◽  
Membrane Translocation ◽  
M3 Receptor

Diverse extracellular signals induce plasma membrane translocation of sphingosine kinase-1 (SphK1), thereby enabling inside-out signaling of sphingosine-1-phosphate. We have shown before that Gq-coupled receptors and constitutively active Gαq/11 specifically induced a rapid and long-lasting SphK1 translocation, independently of canonical Gq/phospholipase C (PLC) signaling. Here, we further characterized Gq/11 regulation of SphK1. SphK1 translocation by the M3 receptor in HEK-293 cells was delayed by expression of catalytically inactive G-protein-coupled receptor kinase-2, p63Rho guanine nucleotide exchange factor (p63RhoGEF), and catalytically inactive PLCβ3, but accelerated by wild-type PLCβ3 and the PLCδ PH domain. Both wild-type SphK1 and catalytically inactive SphK1-G82D reduced M3 receptor-stimulated inositol phosphate production, suggesting competition at Gαq. Embryonic fibroblasts from Gαq/11 double-deficient mice were used to show that amino acids W263 and T257 of Gαq, which interact directly with PLCβ3 and p63RhoGEF, were important for bradykinin B2 receptor-induced SphK1 translocation. Finally, an AIXXPL motif was identified in vertebrate SphK1 (positions 100–105 in human SphK1a), which resembles the Gαq binding motif, ALXXPI, in PLCβ and p63RhoGEF. After M3 receptor stimulation, SphK1-A100E-I101E and SphK1-P104A-L105A translocated in only 25% and 56% of cells, respectively, and translocation efficiency was significantly reduced. The data suggest that both the AIXXPL motif and currently unknown consequences of PLCβ/PLCδ(PH) expression are important for regulation of SphK1 by Gq/11.

scholarly journals Phospholipase C and Protein Kinase C-β 2 Mediate Insulin-Like Growth Factor II-Dependent Sphingosine Kinase 1 Activation

2011 ◽  
Vol 25 (12) ◽  
pp. 2144-2156 ◽  
Author(s):  
Hesham M. El-Shewy ◽  
Souzan A. Abdel-Samie ◽  
Abdelmohsen M. Al Qalam ◽  
Mi-Hye Lee ◽  
Kazuyuki Kitatani ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Fluorescent Protein ◽  
Sphingosine Kinase ◽  
Sphingosine Kinase 1 ◽  
Membrane Translocation ◽  
Kinase C ◽  
Sphingosine 1 Phosphate

Abstract We recently reported that IGF-II binding to the IGF-II/mannose-6-phosphate (M6P) receptor activates the ERK1/2 cascade by triggering sphingosine kinase 1 (SK1)-dependent transactivation of G protein-coupled sphingosine 1-phosphate (S1P) receptors. Here, we investigated the mechanism of IGF-II/M6P receptor-dependent sphingosine kinase 1 (SK1) activation in human embryonic kidney 293 cells. Pretreating cells with protein kinase C (PKC) inhibitor, bisindolylmaleimide-I, abolished IGF-II-stimulated translocation of green fluorescent protein (GFP)-tagged SK1 to the plasma membrane and activation of endogenous SK1, implicating PKC as an upstream regulator of SK1. Using confocal microscopy to examine membrane translocation of GFP-tagged PKCα, β1, β2, δ, and ζ, we found that IGF-II induced rapid, transient, and isoform-specific translocation of GFP-PKCβ2 to the plasma membrane. Immunoblotting of endogenous PKC phosphorylation confirmed PKCβ2 activation in response to IGF-II. Similarly, IGF-II stimulation caused persistent membrane translocation of the kinase-deficient GFP-PKCβ2 (K371R) mutant, which does not dissociate from the membrane after translocation. IGF-II stimulation increased diacylglycerol (DAG) levels, the established activator of classical PKC. Interestingly, the polyunsaturated fraction of DAG was increased, indicating involvement of phosphatidyl inositol/phospholipase C (PLC). Pretreating cells with the PLC inhibitor, U73122, attenuated IGF-II-dependent DAG production and PKCβ2 phosphorylation, blocked membrane translocation of the kinase-deficient GFP-PKCβ2 (K371R) mutant, and reduced sphingosine 1-phosphate production, suggesting that PLC/PKCβ2 are upstream regulators of SK1 in the pathway. Taken together, these data provide evidence that activation of PLC and PKCβ2 by the IGF-II/M6P receptor are required for the activation of SK1.

Gαq-mediated plasma membrane translocation of sphingosine kinase-1 and cross-activation of S1P receptors

2009 ◽  
Vol 1791 (5) ◽  
pp. 357-370 ◽  
Author(s):  
Michael ter Braak ◽  
Kerstin Danneberg ◽  
Karin Lichte ◽  
Kerstin Liphardt ◽  
Nicholas T. Ktistakis ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Sphingosine Kinase ◽  
Sphingosine Kinase 1 ◽  
Membrane Translocation ◽  
S1p Receptors

EGF-and NGF-stimulated translocation of cytohesin-1 to the plasma membrane of PC12 cells requires PI 3-kinase activation and a functional cytohesin-1 PH domain

1999 ◽  
Vol 112 (12) ◽  
pp. 1957-1965 ◽  
Author(s):  
K. Venkateswarlu ◽  
F. Gunn-Moore ◽  
J.M. Tavare ◽  
P.J. Cullen
Keyword(s):  
Plasma Membrane ◽  
Pc12 Cells ◽  
Laser Scanning ◽  
Time Course ◽  
Fluorescent Protein ◽  
Dominant Negative ◽  
Head Group ◽  
Nucleotide Exchange ◽  
Ph Domain

ADP-ribosylation factors (ARFs) are small GTP-binding proteins that function as regulators of eukaryotic vesicle trafficking. Cytohesin-1 is a member of a family of ARF guanine nucleotide-exchange factors that contain a C-terminal pleckstrin homology (PH) domain which has been proposed to bind the lipid second messenger phosphatidylinositol 3,4,5-trisphosphate (PIP3). Here we demonstrate that in vitro, recombinant cytohesin-1 binds, via its PH domain, the inositol head group of PIP3, inositol 1,3,4, 5-tetrakisphosphate (IP4), with an affinity greater than 200-fold higher than the inositol head group of either phosphatidylinositol 4, 5-bisphosphate or phosphatidylinositol 3,4-bisphosphate. Moreover, addition of glycerol or diacetylglycerol to the 1-phosphate of IP4 does not alter the ability to interact with cytohesin-1, data which is entirely consistent with cytohesin-1 functioning as a putative PIP3 receptor. To address whether cytohesin-1 binds PIP3 in vivo, we have expressed a chimera of green fluorescent protein (GFP) fused to the N terminus of cytohesin-1 in PC12 cells. Using laser scanning confocal microscopy we demonstrate that either EGF- or NGF-stimulation of transiently transfected PC12 cells results in a rapid translocation of GFP-cytohesin-1 from the cytosol to the plasma membrane. This translocation is dependent on the cytohesin-1 PH domain and occurs with a time course that parallels the rate of plasma membrane PIP3 production. Furthermore, the translocation requires the ability of either agonist to activate PI 3-kinase, since it is inhibited by wortmannin (100 nM), LY294002 (50 microM) and by coexpression with a dominant negative p85. This data therefore suggests that in vivo cytohesin-1 can interact with PIP3 via its PH domain.

scholarly journals The Calmodulin-binding Site of Sphingosine Kinase and Its Role in Agonist-dependent Translocation of Sphingosine Kinase 1 to the Plasma Membrane

2006 ◽  
Vol 281 (17) ◽  
pp. 11693-11701 ◽  
Author(s):  
Catherine M. Sutherland ◽  
Paul A. B. Moretti ◽  
Niamh M. Hewitt ◽  
Christopher J. Bagley ◽  
Mathew A. Vadas ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Binding Site ◽  
Sphingosine Kinase ◽  
Sphingosine Kinase 1 ◽  
Calmodulin Binding

scholarly journals Active Arf6 Recruits ARNO/Cytohesin GEFs to the PM by Binding Their PH Domains

2007 ◽  
Vol 18 (6) ◽  
pp. 2244-2253 ◽  
Author(s):  
Lee Ann Cohen ◽  
Akira Honda ◽  
Peter Varnai ◽  
Fraser D. Brown ◽  
Tamas Balla ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Family Member ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Ph Domain ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Biochemical Assays ◽  
Inositol Phospholipids ◽  
Sec7 Domain

ARNO is a soluble guanine nucleotide exchange factor (GEF) for the Arf family of GTPases. Although in biochemical assays ARNO prefers Arf1 over Arf6 as a substrate, its localization in cells at the plasma membrane (PM) suggests an interaction with Arf6. In this study, we found that ARNO activated Arf1 in HeLa and COS-7 cells resulting in the recruitment of Arf1 on to dynamic PM ruffles. By contrast, Arf6 was activated less by ARNO than EFA6, a canonical Arf6 GEF. Remarkably, Arf6 in its GTP-bound form recruited ARNO to the PM and the two proteins could be immunoprecipitated. ARNO binding to Arf6 was not mediated through the catalytic Sec7 domain, but via the pleckstrin homology (PH) domain. Active Arf6 also bound the PH domain of Grp1, another ARNO family member. This interaction was direct and required both inositol phospholipids and GTP. We propose a model of sequential Arf activation at the PM whereby Arf6-GTP recruits ARNO family GEFs for further activation of other Arf isoforms.

scholarly journals PKC-dependent Activation of Sphingosine Kinase 1 and Translocation to the Plasma Membrane

2002 ◽  
Vol 277 (38) ◽  
pp. 35257-35262 ◽  
Author(s):  
Korey R. Johnson ◽  
Kevin P. Becker ◽  
Maria Marta Facchinetti ◽  
Yusuf A. Hannun ◽  
Lina M. Obeid
Keyword(s):  
Plasma Membrane ◽  
Sphingosine Kinase ◽  
Sphingosine Kinase 1

scholarly journals Sphingosine kinase-1 and sphingosine 1-phosphate receptor 2 mediate Bcr-Abl1 stability and drug resistance by modulation of protein phosphatase 2A

Blood ◽  
2011 ◽  
Vol 117 (22) ◽  
pp. 5941-5952 ◽  
Author(s):  
Arelis Salas ◽  
Suriyan Ponnusamy ◽  
Can E. Senkal ◽  
Marisa Meyers-Needham ◽  
Shanmugam Panneer Selvam ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Mononuclear Cells ◽  
Blast Crisis ◽  
Chronic Phase ◽  
Sphingosine Kinase ◽  
Sphingosine Kinase 1 ◽  
Wild Type ◽  
Imatinib Resistance ◽  
Novel Approach ◽  
Sphingosine 1 Phosphate

Abstract The mechanisms by which sphingosine kinase-1 (SK-1)/sphingosine 1-phosphate (S1P) activation contributes to imatinib resistance in chronic myeloid leukemia (CML) are unknown. We show herein that increased SK-1/S1P enhances Bcr-Abl1 protein stability, through inhibition of its proteasomal degradation in imatinib-resistant K562/IMA-3 and LAMA-4/IMA human CML cells. In fact, Bcr-Abl1 stability was enhanced by ectopic SK-1 expression. Conversely, siRNA-mediated SK-1 knockdown in K562/IMA-3 cells, or its genetic loss in SK-1−/− MEFs, significantly reduced Bcr-Abl1 stability. Regulation of Bcr-Abl1 by SK-1/S1P was dependent on S1P receptor 2 (S1P2) signaling, which prevented Bcr-Abl1 dephosphorylation, and degradation via inhibition of PP2A. Molecular or pharmacologic interference with SK-1/S1P2 restored PP2A-dependent Bcr-Abl1 dephosphorylation, and enhanced imatinib- or nilotinib-induced growth inhibition in primary CD34+ mononuclear cells obtained from chronic phase and blast crisis CML patients, K562/IMA-3 or LAMA4/IMA cells, and 32Dcl3 murine progenitor cells, expressing the wild-type or mutant (Y253H or T315I) Bcr-Abl1 in situ. Accordingly, impaired SK-1/S1P2 signaling enhanced the growth-inhibitory effects of nilotinib against 32D/T315I-Bcr-Abl1–derived mouse allografts. Since SK-1/S1P/S1P2 signaling regulates Bcr-Abl1 stability via modulation of PP2A, inhibition of SK-1/S1P2 axis represents a novel approach to target wild-type- or mutant-Bcr–Abl1 thereby overcoming drug resistance.

scholarly journals Translocation of Sphingosine Kinase 1 to the Plasma Membrane Is Mediated by Calcium- and Integrin-binding Protein 1

2009 ◽  
Vol 285 (1) ◽  
pp. 483-492 ◽  
Author(s):  
Kate E. Jarman ◽  
Paul A. B. Moretti ◽  
Julia R. Zebol ◽  
Stuart M. Pitson
Keyword(s):  
Plasma Membrane ◽  
Binding Protein ◽  
Sphingosine Kinase ◽  
Sphingosine Kinase 1

scholarly journals Sphingosine and Sphingosine Kinase 1 Involvement in Endocytic Membrane Trafficking

2017 ◽  
Vol 292 (8) ◽  
pp. 3074-3088 ◽  
Author(s):  
Santiago Lima ◽  
Sheldon Milstien ◽  
Sarah Spiegel
Keyword(s):  
Plasma Membrane ◽  
Membrane Trafficking ◽  
Sphingosine Kinase ◽  
Cell Types ◽  
Membrane Cholesterol ◽  
Vesicle Formation ◽  
Sphingosine Kinase 1 ◽  
Late Endosomes ◽  
Sphingosine 1 Phosphate ◽  
Plasma Membrane Cholesterol

The balance between cholesterol and sphingolipids within the plasma membrane has long been implicated in endocytic membrane trafficking. However, in contrast to cholesterol functions, little is still known about the roles of sphingolipids and their metabolites. Perturbing the cholesterol/sphingomyelin balance was shown to induce narrow tubular plasma membrane invaginations enriched with sphingosine kinase 1 (SphK1), the enzyme that converts the bioactive sphingolipid metabolite sphingosine to sphingosine-1-phosphate, and suggested a role for sphingosine phosphorylation in endocytic membrane trafficking. Here we show that sphingosine and sphingosine-like SphK1 inhibitors induced rapid and massive formation of vesicles in diverse cell types that accumulated as dilated late endosomes. However, much smaller vesicles were formed in SphK1-deficient cells. Moreover, inhibition or deletion of SphK1 prolonged the lifetime of sphingosine-induced vesicles. Perturbing the plasma membrane cholesterol/sphingomyelin balance abrogated vesicle formation. This massive endosomal influx was accompanied by dramatic recruitment of the intracellular SphK1 and Bin/Amphiphysin/Rvs domain-containing proteins endophilin-A2 and endophilin-B1 to enlarged endosomes and formation of highly dynamic filamentous networks containing endophilin-B1 and SphK1. Together, our results highlight the importance of sphingosine and its conversion to sphingosine-1-phosphate by SphK1 in endocytic membrane trafficking.

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