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Figure 7 | BMC Developmental Biology

Figure 7

From: The MAP kinase ERK and its scaffold protein MP1 interact with the chromatin regulator Corto during Drosophilawing tissue development

Figure 7

Corto interacts in vivo with ERK in the nucleus only. (A, B): ERK-FLAG co-immunoprecipitates with dMP1-Myc (A) and Corto-Myc (B) only in nuclear extracts. Immunoprecipitation was performed with either anti-FLAG (FLAG-IP) or anti-HA (Mock-IP) antibodies. Cytoplasmic (Cyt) and nuclear (Nuc) extracts from S2 cells expressing tagged proteins were analyzed by Western blot using β-tubulin and lamin as cytoplasmic and nuclear loading controls, respectively. Note that ERK-FLAG (about 50 kDa) co-migrates with the heavy IgG chains (asterisks). (C): Corto co-immunoprecipitates with the di-phosphorylated form of ERK (dP-ERK) in a total embryonic extract. Immunoprecipitated proteins were revealed using rat anti-Corto antibodies. (D): dMP1-Myc co-immunoprecipitates with Corto-FLAG only after cross-linking of cells before protein extraction. Immunoprecipitation was performed using either anti-Myc (Myc-IP) or anti-HA (Mock-IP) antibodies. In A, B, D, immunoprecipitated proteins were revealed by Western-blot using anti-FLAG or anti-Myc antibodies. Arrows and braces show immunoprecipitated tagged proteins and asterisks point to heavy or light IgG chains. In A and D, the light IgG chains in the mock-IP lanes are poorly recognized by the secondary antibodies, but are clearly visible when membranes were stained with Ponceau red (data not shown). S: supernatant after immunoprecipitation; IP: protein G-agarose beads. 5% of the input and 50% of the immunoprecipitate were loaded onto the gel.

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