Fig. 1.

Gtpbp2 is required for Wnt but not nodal target gene induction. a Four-cell Xenopus embryos injected dorsally with 25 ng of Gtpbp2 MO (middle, 93 % abnormal, n = 94), but not a control 5-base mismatch MO (left, 95 % normal, n = 54), generated tadpoles with severe axial defects that were partially rescued by co-injection of the MO and a cocktail of 0.5 ng gtpbp2a and 50 pg gtpbp2b mRNA (right panel, 56 % rescued head and anterior structures, n = 78). b Embryos injected bilaterally at the 2-cell stage with 15 ng Gtpbp2 (gtpbp2 mo) or mismatch control (co-mo) morpholino were cultured until stage 10.25 then measured for nodal3.1, siamois, and ornithine decarboxylase (odc) levels. Amounts of nodal3.1 or siamois were normalized to odc and relative levels shown as mean ± s.e.m of n = 3 with p-values from t-test. c Expression of organizer genes chordin, gsc, and frzb as well as the pan-mesodermal markers t was severely reduced in embryos injected with Gtpbp2 morpholino into two dorsal blastomeres at the four-cell stage. Expression of another T-box gene vegt was not affected. Experiments were repeated three times. d-f mRNAs encoding nodal2 (10 pg; d, e) or wnt8 (10 pg, f) were coinjected with control or Gtpbp2 morpholino into the animal pole of two cell embryos. Animal caps were cut at midblastula (stage 8), and the expression of nodal targets goosecoid (d) and mixer (e), and the Wnt8 target nodal3.1 (f), were measured using qPCR at early gastrula stage 10.5. Relative expression levels shown as mean ± s.e.m (n = 3); p-values from t-test. g Knockdown of Gtpbp2 in caps treated with wnt8 showed that Gtpbp2 is required for induction from a Wnt/β-catenin reporter. Embryos were injected with combinations of 10 pg wnt8, 4 ng GFP, 30 ng Gtpbp2 m1, Gtpbp2 m2 or control morpholinos as indicated, along with 100 pg of Super Topflash and 60 pg of TK-Renilla Luciferase plasmids. Reporter activities were normalized to renilla luciferase activity from a co-injected TK-RL construct and shown as mean ± s.e.m (n = 3) with p-values from t-test