American Society of Hematology

Figure 4.

$Figure 4. Serum induces translocation of SphK1 to the plasma membrane. (A) RBL-2H3 cells stably expressing vector or SphK1 were cultured for 24 hours in the absence (□) or presence (▦) of 15% serum. Cells were homogenized and centrifuged at 1000g to remove unbroken cells and then at 100 000g for 1 hour to obtain cytosol (Cyto) and plasma membrane (PM) fractions. SphK1 activity was measured as described in “Sphingosine kinase assay.” *P < .05; Student t test. (B) Equal amounts of proteins were analyzed by immunoblotting with anti-GFP antibody. (C-D) Duplicate cultures of vector-GFP- or SphK1-GFP-expressing cells were fixed and stained with anti-GFP and examined by confocal fluorescence microscopy (left panels) or by differential interference contrast (DIC; right panels) to visualize the cells.$

Serum induces translocation of SphK1 to the plasma membrane. (A) RBL-2H3 cells stably expressing vector or SphK1 were cultured for 24 hours in the absence (□) or presence (▦) of 15% serum. Cells were homogenized and centrifuged at 1000g to remove unbroken cells and then at 100 000g for 1 hour to obtain cytosol (Cyto) and plasma membrane (PM) fractions. SphK1 activity was measured as described in “Sphingosine kinase assay.” ^*P < .05; Student t test. (B) Equal amounts of proteins were analyzed by immunoblotting with anti-GFP antibody. (C-D) Duplicate cultures of vector-GFP- or SphK1-GFP-expressing cells were fixed and stained with anti-GFP and examined by confocal fluorescence microscopy (left panels) or by differential interference contrast (DIC; right panels) to visualize the cells.

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