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Fig. 1. Antisense MOs against Bmp2, Bmp4 and Bmp7 inhibit endogenous Smad1 phosphorylation and cause dorsalization and posterior truncations of Xenopus embryos. (A) Design of Bmp4, Bmp7 and Bmp2 antisense MOs that target both pseudoalleles expressed in the subtetraploid species X. laevis. (B) In vitro transcription/translation of Bmp4, Bmp7 and Bmp2 is specifically inhibited by the respective MOs. (C) MOs for Bmp2, Bmp4 and Bmp7 (at 12 ng each) were injected either alone or in combinations at the four-cell stage radially in each blastomere. (D) Endogenous carboxy-terminal Smad1 phosphorylation in stage 11 embryos is decreased by co-injection of multiple Bmp MOs. (E-H) Bmp4 MO-injected embryos (F) are dorsalized with enlarged heads (compare with control embryos, E). Red arrowheads delineate the spinal cord marker Hoxb9 (>85%, n=60). (G) Bmp4 MO-dorsalized phenotype is rescued by microinjection of 100 pg of mouse Bmp4 mRNA. (H) Microinjection of mouse Bmp4 mRNA alone (100 pg) results in ventralized embryos with small heads, no eyes and reduced spinal cord structures (red arrowheads). (I-L) Bmp4-depleted embryos (J) develop into swimming tadpoles with no ventral fins and slightly larger heads than control embryos (I). Note the position of the anus, which is displaced (posteriorized) to the tip of the tail (white arrowhead; >72%, n=61). (K) Bmp7-depleted tadpoles develop with a partial loss of ventral fin and a posteriorized anus (white arrowhead;> 79%, n=51). (L) Double knockdown of Bmp4 and Bmp7 results in tadpoles lacking tail structures (>90%, n=39). |
