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FIGURE 9. Evidence that XSMOC-1 signals through the MAPK pathway. A, XSMOC-1 activity was blocked by co-expression of LM-Smad1. RT-PCR anal- ysis of animal caps from embryos injected bilaterally at the two-cell stage with 300 pg of XSMOC-1 plus 600 pg of GFP, 300 pg of XSMOC-1 plus 600 pg of LM-Smad1, 6 pg of noggin plus 900 pg of GFP, or 6 pg of noggin plus 600 pg of LM-Smad1 and 300 pg of GFP mRNAs. Induction of the neural markers N-CAM, NRP1, and Otx2 by overexpression of XSMOC-1 was blocked by co- expression of LM-Smad1; expression of the epidermal marker keratin was maintained. Neural marker induction (and suppression of keratin) by over-expression of noggin was not affected by co-expression of LM-Smad1. B, immunoblot analysis of animal cap extracts from embryos over-eexpressing XSMOC-1 revealed elevated levels of dp-ERK. Equivalent amounts of protein (10 mg) were loaded per lane. C, anterior views of control (left) and XSMOC-1 MO-injected stage 12.5 embryos immunostained for dp-ERK, Note the absence of dp-ERK in the XSMOC-1 MO-injected embryo. D, RT-PCR analysis of animal caps from XSMOC-1-injected embryos incubated in the presence or absence of the MEK inhibitor U0126 (50uM) until control embryos reached stage 17. Anterior neuroectodermal (Otx2 and XAG-1) and panneural (NCAM and NRP-1) markers induced by XSMOC-1 were markedly reduced in the pres- ence of U0126. -RT, without reverse transcriptase.
""This research was originally published THE JOURNAL OF BIOLOGICAL CHEMISTRY VOL.284(28) pp.18994-19005, July 10, 2009. Copyright: The American Society for Biochemistry and Molecular Biology." |
