Soluble klotho binds monosialoganglioside to regulate membrane microdomains and growth factor signaling

Significance Soluble klotho is the shed ectodomain of the antiaging membrane protein α-klotho that exhibits pleiotropic actions, including down-regulation of growth factor-driven PI3K signaling, contributing to lifespan prolongation, cardioprotection, and tumor inhibition. Whether membrane receptors exist for soluble klotho is unknown. We identify lipid rafts as receptors for soluble klotho. We show klotho binds specific sialic acid residues of gangliosides highly enriched in the outer leaflet of lipid rafts. Klotho binding to gangliosides modulates lipid organization and inhibits lipid raft-dependent PI3K signaling. In vivo, klotho-deficient mouse hearts have heightened raft-dependent PI3K signaling vs. wild-type hearts. We reveal a novel physiological regulator of lipid raft structure and function, and open new research to understand pleiotropic effects of klotho in cell signaling and metabolism. Soluble klotho, the shed ectodomain of the antiaging membrane protein α-klotho, is a pleiotropic endocrine/paracrine factor with no known receptors and poorly understood mechanism of action. Soluble klotho down-regulates growth factor-driven PI3K signaling, contributing to extension of lifespan, cardioprotection, and tumor inhibition. Here we show that soluble klotho binds membrane lipid rafts. Klotho binding to rafts alters lipid organization, decreases membrane’s propensity to form large ordered domains for endocytosis, and down-regulates raft-dependent PI3K/Akt signaling. We identify α2-3-sialyllactose present in the glycan of monosialogangliosides as targets of soluble klotho. α2-3-Sialyllactose is a common motif of glycans. To explain why klotho preferentially targets lipid rafts we show that clustering of gangliosides in lipid rafts is important. In vivo, raft-dependent PI3K signaling is up-regulated in klotho-deficient mouse hearts vs. wild-type hearts. Our results identify ganglioside-enriched lipid rafts to be receptors that mediate soluble klotho regulation of PI3K signaling. Targeting sialic acids may be a general mechanism for pleiotropic actions of soluble klotho.

[1]  S. Houser,et al.  Transient Receptor Potential Channels Contribute to Pathological Structural and Functional Remodeling After Myocardial Infarction , 2014, Circulation research.

[2]  C. Abraham,et al.  Biochemical and Functional Characterization of the Klotho-VS Polymorphism Implicated in Aging and Disease Risk* , 2013, The Journal of Biological Chemistry.

[3]  Youhua Liu,et al.  Loss of Klotho contributes to kidney injury by derepression of Wnt/β-catenin signaling. , 2013, Journal of the American Society of Nephrology : JASN.

[4]  T. Pawson,et al.  Soluble FLT1 Binds Lipid Microdomains in Podocytes to Control Cell Morphology and Glomerular Barrier Function , 2012, Cell.

[5]  M. Rao,et al.  Active Remodeling of Cortical Actin Regulates Spatiotemporal Organization of Cell Surface Molecules , 2012, Cell.

[6]  J. Xie,et al.  Cardioprotection by Klotho through downregulation of TRPC6 channels in the mouse heart , 2012, Nature Communications.

[7]  K. Gaus,et al.  Quantitative imaging of membrane lipid order in cells and organisms , 2011, Nature Protocols.

[8]  Kai Simons,et al.  Membrane organization and lipid rafts. , 2011, Cold Spring Harbor perspectives in biology.

[9]  Xin Zhou,et al.  PI3K/Akt signaling requires spatial compartmentalization in plasma membrane microdomains , 2011, Proceedings of the National Academy of Sciences.

[10]  Chou-Long Huang,et al.  Klotho: a novel regulator of calcium and phosphorus homeostasis , 2011, Pflügers Archiv - European Journal of Physiology.

[11]  D. Hilgemann,et al.  Mechanistic analysis of massive endocytosis in relation to functionally defined surface membrane domains , 2011, The Journal of general physiology.

[12]  D. Lingwood,et al.  Lipid rafts as functional heterogeneity in cell membranes. , 2009, Biochemical Society transactions.

[13]  P. Matarrese,et al.  Fas death receptor enhances endocytic membrane traffic converging into the Golgi region. , 2008, Molecular biology of the cell.

[14]  K. Rosenblatt,et al.  Removal of sialic acid involving Klotho causes cell-surface retention of TRPV5 channel via binding to galectin-1 , 2008, Proceedings of the National Academy of Sciences.

[15]  E. Gratton,et al.  The phasor approach to fluorescence lifetime imaging analysis. , 2008, Biophysical journal.

[16]  C. Kuo,et al.  Augmented Wnt Signaling in a Mammalian Model of Accelerated Aging , 2007, Science.

[17]  M. Neil,et al.  Fluorescence lifetime imaging provides enhanced contrast when imaging the phase-sensitive dye di-4-ANEPPDHQ in model membranes and live cells. , 2006, Biophysical journal.

[18]  K. Rosenblatt,et al.  Regulation of Fibroblast Growth Factor-23 Signaling by Klotho* , 2006, Journal of Biological Chemistry.

[19]  J. Hoenderop,et al.  The ß-Glucuronidase Klotho Hydrolyzes and Activates the TRPV5 Channel , 2005, Science.

[20]  Animesh Nandi,et al.  Suppression of Aging in Mice by the Hormone Klotho , 2005, Science.

[21]  K. Nozaki,et al.  Secreted Klotho protein in sera and CSF: implication for post‐translational cleavage in release of Klotho protein from cell membrane , 2004, FEBS letters.

[22]  M. Lussier,et al.  Exocytotic Insertion of TRPC6 Channel into the Plasma Membrane upon Gq Protein-coupled Receptor Activation* , 2004, Journal of Biological Chemistry.

[23]  Y. Hayashizaki,et al.  Identification of a novel mouse membrane-bound family 1 glycosidase-like protein, which carries an atypical active site structure. , 2002, Biochimica et biophysica acta.

[24]  R. Dwek,et al.  N-butyldeoxygalactonojirimycin: a more selective inhibitor of glycosphingolipid biosynthesis than N-butyldeoxynojirimycin, in vitro and in vivo. , 2000, Biochemical pharmacology.

[25]  T. Gudermann,et al.  Direct activation of human TRPC6 and TRPC3 channels by diacylglycerol , 1999, Nature.

[26]  A. May,et al.  Crystal structure of the N-terminal domain of sialoadhesin in complex with 3' sialyllactose at 1.85 A resolution. , 1998, Molecular cell.

[27]  Tadashi Kaname,et al.  Mutation of the mouse klotho gene leads to a syndrome resembling ageing , 1997, Nature.

[28]  J. Xie,et al.  Klotho May Ameliorate Proteinuria by Targeting TRPC6 Channels in Podocytes. , 2017, Journal of the American Society of Nephrology : JASN.

[29]  A. Varki,et al.  Modulation of glycan recognition by clustered saccharide patches. , 2014, International review of cell and molecular biology.

[30]  J. Hsuan,et al.  Preparation of membrane rafts. , 2009, Methods in molecular biology.

[31]  M. Čačić,et al.  Glycosphingolipid expression in human skeletal and heart muscle assessed by immunostaining thin-layer chromatography , 2004, Glycoconjugate Journal.