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Figure 6 | BMC Developmental Biology

Figure 6

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

Figure 6

The phosphorylation pattern of Corto depends on the level of ERK pathway activity. (A): S2 cells in 0% fetal calf serum were transfected with a plasmid allowing Corto-FLAG over-expression. 48 hours after transfection, they were submitted to short ERK pathway activation by a 15 minutes treatment with either 10% fetal calf serum (lane 2), 10% fetal calf serum plus insulin 10 μg/ml (lane 3) or 10% fetal calf serum plus PMA 100 ng/ml (lane 4). Total protein extracts were either directly analyzed by Western blot with anti-FLAG antibodies (upper panel), or submitted to immunoprecipitation using anti-FLAG antibodies followed by immunoblotting with anti-phosphoprotein antibodies (lower panel). The braces point to the upper Corto isoforms that appear in response to ERK activation. The asterisk on the lower panel shows the heavy IgG chains, which have already been shown to be phosphorylated [62]. (B): Serum-starved S2 cells were either transfected with a plasmid allowing Corto-FLAG expression (lane 1), or co-transfected with Corto-FLAG and ERK-FLAG (lane 2), Corto-FLAG and ERKSem-FLAG (lane 3) or Corto-FLAG and RasV12 (lane 4), in order to constitutively activate the ERK pathway. After 48 hours, total protein extracts were analyzed by Western blot using anti-FLAG antibodies.

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