Fig. 2.
Fig. 2. Use of antisense oligonucleotides to reduce specific expression of either PLD1 or PLD2 demonstrates that only PLD1 is coupled to FcγRI aggregation. / (A) Western blot analysis of immunoprecipitates of either PLD1 or PLD2 to assess expression of either isozyme in IFN-γ–primed U937 cells following treatment for 36 hours with antisense oligonucleotides (10 μM) specific for either PLD1 (a.s.PLD1) or PLD2 (a.s.PLD2), and control cells (control). The results shown are typical from 3 separate experiments. (B) PLD activity following FcγRI aggregation in IFN-γ–primed U937 cells pretreated with 10 μM antisense oligonucleotides for either PLD1 (a.s.PLD1) or PLD2 (a.s.PLD2). 1. Basal level (basal control); 2. FcγRI aggregation (XL control); 3. basal level in cells pretreated with antisense PLD1 (basal a.s.PLD1); 4. FcγRI aggregation in cells pretreated with antisense PLD1 (XL a.s.PLD1); 5. basal level in cells pretreated with antisense PLD2 (basal a.s.PLD2); 6. FcγRI aggregation in cells pretreated with antisense PLD2 (XL a.s.PLD2). Results are the mean ± SD for triplicate measurements and are representative of the results from 3 separate experiments. PtdBut indicates phosphatidylbutanol. (C) PLD activity following PMA stimulation (1 μM) in IFN-γ–primed U937 cells pretreated with antisense oligonucleotides (10 μM) for either PLD1 (a.s.PLD1) or PLD2 (a.s.PLD2). 1. Basal level (basal control); 2. PMA stimulation (PMA control); 3. basal level in cells pretreated with antisense PLD1 (basal a.s.PLD1); 4. PMA stimulation in cells pretreated with antisense PLD1 (PMA a.s.PLD1); 5. basal level in cells pretreated with antisense PLD2 (basal a.s.PLD2); 6. PMA stimulation in cells pretreated with antisense PLD2 (PMA a.s.PLD2). Results are the mean ± SD for triplicate measurements and are representative of the results from at least 3 separate experiments. Tot. indicates total.

Use of antisense oligonucleotides to reduce specific expression of either PLD1 or PLD2 demonstrates that only PLD1 is coupled to FcγRI aggregation.

(A) Western blot analysis of immunoprecipitates of either PLD1 or PLD2 to assess expression of either isozyme in IFN-γ–primed U937 cells following treatment for 36 hours with antisense oligonucleotides (10 μM) specific for either PLD1 (a.s.PLD1) or PLD2 (a.s.PLD2), and control cells (control). The results shown are typical from 3 separate experiments. (B) PLD activity following FcγRI aggregation in IFN-γ–primed U937 cells pretreated with 10 μM antisense oligonucleotides for either PLD1 (a.s.PLD1) or PLD2 (a.s.PLD2). 1. Basal level (basal control); 2. FcγRI aggregation (XL control); 3. basal level in cells pretreated with antisense PLD1 (basal a.s.PLD1); 4. FcγRI aggregation in cells pretreated with antisense PLD1 (XL a.s.PLD1); 5. basal level in cells pretreated with antisense PLD2 (basal a.s.PLD2); 6. FcγRI aggregation in cells pretreated with antisense PLD2 (XL a.s.PLD2). Results are the mean ± SD for triplicate measurements and are representative of the results from 3 separate experiments. PtdBut indicates phosphatidylbutanol. (C) PLD activity following PMA stimulation (1 μM) in IFN-γ–primed U937 cells pretreated with antisense oligonucleotides (10 μM) for either PLD1 (a.s.PLD1) or PLD2 (a.s.PLD2). 1. Basal level (basal control); 2. PMA stimulation (PMA control); 3. basal level in cells pretreated with antisense PLD1 (basal a.s.PLD1); 4. PMA stimulation in cells pretreated with antisense PLD1 (PMA a.s.PLD1); 5. basal level in cells pretreated with antisense PLD2 (basal a.s.PLD2); 6. PMA stimulation in cells pretreated with antisense PLD2 (PMA a.s.PLD2). Results are the mean ± SD for triplicate measurements and are representative of the results from at least 3 separate experiments. Tot. indicates total.

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