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Fig. 4. DUSP6 is a negative feedback regulator of the ERK pathway. (A-C) Immunostaining of wild-type and Dusp6^-/- embryos with anti-dpERK. (A) An E9.5 Dusp6^-/- embryo has increased levels of dpERK in the limb (black arrow) relative to that of a wild-type embryo (white arrow). (B) Limb bud of wild-type E10.5 embryo. (C) Limb bud of Dusp6^-/- embryo with increased levels of dpERK. (D-F) RNA in situ hybridization of E10.5 embryos shows increasing levels of Erm transcripts as Dusp6 levels decrease. Genotype of each embryo is indicated in the lower left of each panel. Northern blot hybridization of mRNA isolated from pooled E11.5 embryos (G) or individual adult brains (H) of the indicated genotypes and probed sequentially with a Dusp6 5' UTR probe, a LacZ probe and Gapdh shows that embryos lacking Dusp6 have increased levels of transcripts initiated from the Dusp6 promoter. Note that the embryonic Gapdh panel is identical to the one shown in Fig. 3E, because it comes from the same blot. (I) Diagrams of the transcripts produced by wild-type (Dusp6⁺) and mutant (Dusp6^L) alleles and the locations of the probes (bars above transcripts) used for hybridization. Open boxes, untranslated Dusp6 sequence; black boxes, DUSP6 coding sequence; gray box, LacZ cassette.

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