American Society of Hematology

Figure 2.

$Figure 2. Immunoprecipitations analysis of the mb-IL-15 present on human spleen myofibroblasts. (A) Coimmunoprecipitation with an anti-IL-15 on plasma-membrane fraction. The membrane was then subsequently reprobed by Western blotting (WB) with anti-IL-15Rα, β, γc, or anti-IL-15 Ab: a single 56-kDa specific band was detected by anti-IL-15Rα and a single 14-kDa band with anti-IL-15 Ab. The anti-IL-15Rβ and γc Ab detected specific bands at 70 and 64 kDa, respectively. The isotype control (anti-GM-CSFRβ chain Ab) did not detect any band. (B) Coimmunoprecipitation with an anti-IL-15 on brefeldin A-treated cytosolic fraction. The membrane was then subsequently reprobed by Western blotting with anti-IL-15Rα, β, γc, or anti-IL-15 Ab: a single 14-kDa specific band was detected by anti-IL-15 and a single 56-kDa band with anti-IL-15Rα Ab. No specific bands were detected with the anti-IL-15Rβ and γc Ab. The isotype control (anti-GM-CSFRβ chain Ab) did not detect any band. These results are representative of 3 different experiments. Our data suggest that an IL-15/IL-15Rα complex is assembled intracellularly (B) and then migrates to the cellular membrane, where it associates with the IL-15Rβγc chains (A).$

Immunoprecipitations analysis of the mb-IL-15 present on human spleen myofibroblasts. (A) Coimmunoprecipitation with an anti-IL-15 on plasma-membrane fraction. The membrane was then subsequently reprobed by Western blotting (WB) with anti-IL-15Rα, β, γc, or anti-IL-15 Ab: a single 56-kDa specific band was detected by anti-IL-15Rα and a single 14-kDa band with anti-IL-15 Ab. The anti-IL-15Rβ and γc Ab detected specific bands at 70 and 64 kDa, respectively. The isotype control (anti-GM-CSFRβ chain Ab) did not detect any band. (B) Coimmunoprecipitation with an anti-IL-15 on brefeldin A-treated cytosolic fraction. The membrane was then subsequently reprobed by Western blotting with anti-IL-15Rα, β, γc, or anti-IL-15 Ab: a single 14-kDa specific band was detected by anti-IL-15 and a single 56-kDa band with anti-IL-15Rα Ab. No specific bands were detected with the anti-IL-15Rβ and γc Ab. The isotype control (anti-GM-CSFRβ chain Ab) did not detect any band. These results are representative of 3 different experiments. Our data suggest that an IL-15/IL-15Rα complex is assembled intracellularly (B) and then migrates to the cellular membrane, where it associates with the IL-15Rβγc chains (A).

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