Dissecting [G.sub.q/11]-Mediated Plasma Membrane Translocation of Sphingosine Kinase-1

Diverse extracellular signals induce plasma membrane translocation of sphingosine kinase-1 (SphKl), thereby enabling inside-out signaling of sphingosine-l-phosphate. We have shown before that [G.sub.q]-coupled receptors and constitutively active [G[alpha].sub.q/11] specifically induced a rapid and l...

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Published in:Cells (Basel, Switzerland) Switzerland), 2020-10, Vol.9 (10), p.1
Main Authors: Blankenbach, Kira Vanessa, Claas, Ralf Frederik, Aster, Natalie Judith, Spohner, Anna Katharina, Trautmann, Sandra, Ferreiros, Nerea, Black, Justin L, Tesmer, John J.G, Offermanns, Stefan, Wieland, Thomas, zu Heringdorf, Dagmar Meyer
Format: Article
Language:English
Subjects:
Cell membranes
Phosphotransferases
Physiological aspects
Sphingosine
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Summary:Diverse extracellular signals induce plasma membrane translocation of sphingosine kinase-1 (SphKl), thereby enabling inside-out signaling of sphingosine-l-phosphate. We have shown before that [G.sub.q]-coupled receptors and constitutively active [G[alpha].sub.q/11] specifically induced a rapid and long-lasting SphKl translocation, independently of canonical [G.sub.q]/phospholipase C (PLC) signaling. Here, we further characterized [G.sub.q/11] regulation of SphKl. SphKl translocation by the [M.sub.3] 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 PLC[[beta].sub.3], but accelerated by wild-type PLC[[beta].sub.3] and the PLC[delta] PH domain. Both wild-type SphKl and catalytically inactive SphKl-G82D reduced [M.sub.3] receptor-stimulated inositol phosphate production, suggesting competition at [G[alpha].sub.q]. Embryonic fibroblasts from [G[alpha].sub.q/11] double-deficient mice were used to show that amino acids W263 and T257 of [G[alpha].sub.q], which interact directly with PLC[[beta].sub.3] and p63RhoGEF, were important for bradykinin [B.sub.2] receptor-induced SphKl translocation. Finally, an AIXXPL motif was identified in vertebrate SphKl (positions 100-105 in human SphKla), which resembles the [G[alpha].sub.q] binding motif, ALXXPI, in PLC[beta] and p63RhoGEF. After [M.sub.3] receptor stimulation, SphKl-AlOOE-IlOlE and SphKl-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 PLC[beta]/PLC[delta](PH) expression are important for regulation of SphKl by [G.sub.q/11].
ISSN:2073-4409
2073-4409
DOI:10.3390/cells9102201