Figure 7
SDF-1α -induced chemoataxis in Jurkat T cells is dependent on PTX-sensitive G i proteins and PKD2. (A) Expression of PLCβ2/3 in Jurkat T cells were examined alongside with parental HEK293 cells or HEK293 cells overexpressing PLCβ2/3. (B) Bar diagram showing maximum SDF-1α-induced Ca2+ mobilization in Jurkat T cells with or without PTX pretreatment. (C) Jurkat T cells pretreated with or without PTX were subjected to SDF-1α-induced chemotactic assay. The chemotactic index was expressed as the ratio of the numbers of cells found in the lower compartments, between the agonist-stimulated and the unstimulated groups. (D) Jurkat T cells were pretreated with or without specific inhibitors for PKD (100 nM Gö6976), PI3K (100 nM wortmannin), and MEK/ERK (10 μM U0126) and then subjected to the chemotactic assay in the presence of SDF-1α. (E) Jurkat T cells pretreated with or without PTX were stimulated with 10 nM SDF-1α for the indicated durations. Cell lysates were subjected to immunoblotting for ERK and PKD2 phosphorylation. (F) Jurkat T cells were transfected with vehicle (RNase free water), scrambled siRNA (siCon), siPKD1, siPKD2 or siPKD3 oligonucleotides for 72 h prior to chemotactic assay. Cells were also harvested and lysates were subjected to Western blot analysis with specific antibody against PKD2. β-actin was used as loading control.