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Fig. S4 Irx7 and Meis1.1 do not interact in vitro. A) Detection of Irx7 in zebrafish embryos by the Irx7 antibody. Nuclear protein extracts from 100% epiboly zebrafish embryos were submitted to Western-blot performed with the Irx7 antibody, or with the preimmune serum. In uninjected embryos (ni), Irx7 is detected by the antibody with an apparent molecular weight of 43 kDa, but not by the preimmune serum. The specificity of the antibody is confirmed by the absence of a band in Western blots performed on Mo7 injected embryos. B) Coimmunoprecipitations of a HA-tagged Irx7 protein and a Myc-tagged Meis1.1 protein, produced in transfected COS7 cells. Irx7HA and Meis1.1Myc were mixed and immunoprecipitated with antibodies specific for either Irx7 or Myc. The precipitation products were assayed by Western-blotting for Irx7 (upper panel) and Myc (lower panel). Irx7 and Meis1.1 do not coimmunoprecipitate. C) Anti-Myc coimmunoprecipitation of in vitro translated proteins, using the reticulocyte lysate system. Irx7 was translated together with Pbx4Myc (lanes 1 and 2), Meis1.1Myc and Pbx4 (lanes 3 and 4), or Meis1.1Myc and LacZ as a control (lanes 5 and 6) in the presence of 35Smethionine, and analysed by autoradiography either directly or following an anti-myc co-immunoprecipitation. Pbx4 protein coimmunoprecipitated with Meis1.1Myc (black rectangle) whereas Irx7 was not coimmunoprecipitated. Note that the Irx7 protein produced by in vitro translation is present as two bands. Ab: antibody; preI: preimmune serum.
Reprinted from Developmental Biology, 327(2), Stedman, A., Lecaudey, V., Havis, E., Anselme, I., Wassef, M., Gilardi-Hebenstreit, P., and Schneider-Maunoury, S., A functional interaction between Irx and Meis patterns the anterior hindbrain and activates krox20 expression in rhombomere 3, 566-577, Copyright (2009) with permission from Elsevier. Full text @ Dev. Biol.