Figure 1

Spn-F interacts physically with Ik2 and the C-terminus of Spn-F is crucial for this interaction. (A) Co-immunoprecipitation of GFP-Ik2 with HA-Spn-F. Proteins were extracted from S2 cells co-transfected with GFP-Ik2 and HA-Spn-F. Untreated lysate was run on gel (Total). The same lysate was incubated with α-GFP antibody and precipitated by protein A-coated beads (IP-α-GFP), or was precipitated by protein A-coated beads without anti-GFP antibody (control). To detect Spn-F, western blotting was performed using α-HA antibodies. HA-Spn-F was successfully precipitated with GFP-Ik2. (B) Co-immunoprecipitation of GFP-Ik2 with HA-Spn-F in the germline. Proteins were extracted from transgenic fly ovaries expressing either GFP-Ik2 with HA-Spn-F (right panel) or HA-Spn-F (control, left panel) in the germline. Ovarian extracts were incubated with α-GFP antibody and precipitated by protein A-coated beads (IP-α-GFP). To detect Spn-F western blotting was performed using α-HA antibodies. HA-Spn-F was successfully precipitated with GFP-Ik2. (C) Schematic diagram of Spn-F protein coil-coiled domains and truncated protein used in D. (D) Co-immunoprecipitation assay of myc-Ik2 with GFP-N'-Spn-F or GFP-C'-Spn-F. Proteins were extracted from S2 cells co-transfected with myc-Ik2 and GFP-N'-Spn-F or GFP-C'-Spn-F. Untreated lysate was run on gel (Total). The same lysate was incubated with α-GFP antibody and precipitated by protein A-coated beads (IP-α-GFP), or was precipitated by protein A-coated beads without anti-GFP antibody (control). To detect Ik2, western blotting was performed using α-myc antibodies. Myc-Ik2 was co-precipitated with the C-terminus of Spn-F but not with the N-terminus of Spn-F.