Journal: Nature Communications

Article Title: Sphingosine-1-phosphate promotes erythrocyte glycolysis and oxygen release for adaptation to high-altitude hypoxia

doi: 10.1038/ncomms12086

Figure Lengend Snippet: ( a ) Pull-down of Hb by lysophosphatic acid, sphingosine and S1P beads from normal human RBC lysates. ( b ) Membrane heme concentrations in isolated Sphk1 −/− mouse erythrocyte pretreated with DMSO, 2 and 6 μmol l −1 S1P in normoxia and hypoxia (4% O 2 ) for 6 h. ( c ) Schematic drawing illustrates functional experiments to monitor translocalization of GAPDH from membrane isolated from human erythrocytes. Hb binding to membrane ( d ) and GAPDH release from membrane to the cytosol ( e ) in human erythrocyte membrane ghost treated with Hb and different concentrations of S1P under normoxia and hypoxia. Hb binding to membrane ( f ) and GAPDH release from membrane to the cytosol ( g ) in human erythrocyte membrane ghost treated with Hb-CO and different concentrations of S1P under normoxia and hypoxia. ( h ) Working model: hypoxia-mediated elevation of erythrocyte Sphk1 activity increases the level of S1P, which binds to deoxy-Hb and facilitates binding of deoxy-Hb to membrane and release of GAPDH; increased cytosolic GAPDH accelerates glycolysis and shifts glucose metabolism in favour of 2,3-BPG production, which in turn leads to more O 2 release to counteract tissue hypoxia. Mean±s.e.m; n =6 per group, * P <0.05 versus normoxia, ** P <0.05 versus 2 μmol l −1 or 100 nmol l −1 , Student's t -test and one-way ANOVA.

Article Snippet: Approximately 100 μl of various lipids conjugated to agarose beads including S1P-agarose beads, lysophosphatic acid beads or sphingosine beads (Echelon Biosciences Inc., Salt Lake City, UT) were washed twice with lysis buffer.

Techniques: Isolation, Functional Assay, Binding Assay, Activity Assay