Siamois and Twin are redundant and essential in formation of the Spemann organizer - PubMed

Fig. 6

Siamois and Twin function redundantly in axial development and organizer formation. (A-L) At the 4-cell stage both dorsal blastomeres were injected with (B, D) a non-specific control morpholino oligonucleotide (NSMO, 50ng), (E,G) a Sia-specific oligonucleotide (SiaMO, 25ng), (F, H) a Twn-specific oligonucleotide (TwnMO, 25ng), or (I, K) a lower dose combination of the Sia and Twn oligonucleotides (SiaMO+TwnMO, 25ng+25ng) or (J, L) a higher dose combination of Sia and Twn oligonucleotides (SiaMO+TwnMO, 50ng+50ng). (A-B, E-F, I-J) Whole embryo morphology (dorsal up, anterior right) and (C-D, G-H, K-L) transverse histological sections (dorsal up) are shown at the tailbud stage. Dorsoanterior index (DAI) is indicated in the lower right corner for these representative embryos. (M) Quantification of axial defects (DAI scores) observed for Control, NSMO, SiaMO, TwnMO and increasing doses of Sia+TwnMO. Axial structures are indicated for the histological sections (n, notochord; sm, somitic muscle; nt, neural tube). Axial development was normal for embryos injected with the individual control, Sia or Twn MO, while coinjection of Sia and Twn MO resulted in severe axial defects, including loss of head structures, and reduction or loss of trunk and tail structures. Histological samples are presented for two examples of the double knockdown phenotype; (K) a partial loss of axial development with absence of notochord, somitic muscle crossing the midline, and mispatterning of the neural tube, and (L) a complete loss of axial development with no notochord, somitic muscle or neural tube. (N-L’) Whole mount in situ hybridization analysis of gene expression at the early gastrula stage (stage 10.25). Embryos injected with 50ng each of NSMO (S-W), SiaMO (X-B’), TwnMO (C’-G’) or a combination of SiaMO and TwnMO (50ng + 50ng) (H’-L’) were analyzed for organizer expression of Gsc (N, S, X, C’, H’) and Chordin (O, T, Y, D’, I’), ventrolateral expression of Xwnt8 (P, U, Z, E’, J’), panmesodermal expression of Xbra (Q, V, A’, F’, K’), and neural plate expression of Opal (R, W, B’, G’, L’). Shown are vegetal views with dorsal up (Gsc, Chordin, Xwnt8 and Xbra) and dorsal-vegetal views with dorsal up (Opal). Double knockdown of Sia and Twn together resulted in a reduction or loss of Gsc expression in 77% of embryos and a reduction or loss of Chordin expression in 100% of embryos.(A,C,N-R) Uninjected control embryos.