XB-ART-2256Dev Cell. March 1, 2005; 8 (3): 401-11.
Depletion of three BMP antagonists from Spemann''s organizer leads to a catastrophic loss of dorsal structures.
Transplanted Spemann''s organizer induces dorsal embryonic cell fates such as the nervous system and somites, but in normal development, elimination of individual organizer signals (such as the bone morphogenetic protein [BMP] antagonists) has surprisingly modest effects on these tissues. Thus, the role of BMP antagonists may be limited to fine tuning the size of the dorsal domain. However, at least five BMP antagonists are specifically expressed in the organizer, and all can mimic aspects of organizer function, suggesting overlapping functions. Here, we deplete the function of three BMP antagonists, chordin, noggin, and follistatin, in Xenopus tropicalis. We demonstrate that this results in catastrophic failure of dorsal development and expansion of ventral and posterior fates. We conclude that BMP antagonists are required for formation of the neural plate and dorsal mesoderm. In addition, our results show that neural specification requires the continuous activity of BMP antagonists from blastula through gastrula stages.
PubMed ID: 15737935
Article link: Dev Cell.
Grant support: GM49346 NIGMS NIH HHS , GM66684 NIGMS NIH HHS , K08-HD42550 NICHD NIH HHS , K12-HD00850 NICHD NIH HHS
Genes referenced: bambi bmp4 cer1 chrd.1 fst gsc msx1 myf5 myod1 nodal3.1 nog not shh sia1 sox2 sox3 szl tbxt ventx2.2 xk81a1
Morpholinos referenced: chrd MO7 fst MO3 nog MO2
Article Images: [+] show captions
|Figure 1. BMP Antagonists Are Expressed in Spemann’s OrganizerBlastula and gastrula embryos were bisected, stained, and then cleared to visualize expression of follistatin (A, F, K, and P), chordin (B, G, L, and Q), noggin (C, H, M, and R), xnr3 (D, I, N, and S), and cerberus (E, J, O, and T) at midblastula (A–E), late blastula (F–J), early gastrula (K–O), and midgastrula (P–T). Dorsal is to the right with the animal pole toward the top of the figure. St = stage.|
|Figure 2. Single Morphants and Double Morphants Still Form a Substantial Neural PlateAll panels show sox2 expression to visualize the neural plate (dorsal views with anterior to the bottom) in neurula embryos (st 14–15). Uninjected embryos are shown (A, D, and G) with sibling noggin morphants (B), follistatin morphants (E), and chordin morphants (H). Double morphants include follistatin/noggin (K), chordin/noggin (N), and chordin/follistatin (Q) and are shown with sibling uninjected embryos (J, M, and P). Morphants were rescued with pufferfish noggin (C, F, I, L, O, and R). Abbreviations are as follows: F = follistatin, C = chordin, N = noggin, UC = uninjected sibling control embryos, MO = morphant, and MO + R = morphant rescued with noggin mRNA.|
|Figure 3. FCN Morphants Have a Catastrophic Loss in Dorsal DevelopmentAll panels are dorsal views with anterior to the top of st 14–15 neurula embryos. Expression of multiple dorsal markers (sox2 [A–D], sox3 [E–H], myf5 [I–L], myoD [M–P], shh [Q–T], and xnot [U–X]) are shown in uninjected sibling embryos (A, C, E, G, I, K, M, O, Q, S, U, and W) and follistatin, chordin, and noggin (FCN) triple morphants (B, F, J, N, R, and V) as well as β-catenin morphants (D, H, L, P, T, and X) for comparison.|
|Figure 4. The Phenotype of the FCN Triple Morphant Is Specific to a Loss in BMP AntagonismAll panels are dorsal views of neurula embryos (st 14) with anterior to the top. Triple morphants were subsequently injected with pufferfish noggin mRNA (C, F, and I) and show substantial rescue of dorsal structures (sox2 [A–C], myoD [D–F], and shh [G–I]) when compared to triple morphants (B, E, and H) and uninjected sibling embryos (A, D, and G). The distribution of phenotypes seen in these rescue experiments is depicted in a bar graph (J). The vertical axis is the percent of embryos that show a particular phenotype. The horizontal axis shows the experimental groups and the phenotypes seen. The black bar depicts the percentage of embryos with a substantial neural plate, the dark gray bar depicts the percentage of embryos with a minor neural plate, and the light gray bar depicts the percentage of embryos with no neural plate that show only a ring of sox2 expression. We also tested the phenotype of the triple morphant for specificity by subsequently injecting BMP4,7 MOs (BMP MO), which should reduce BMP signaling. A partial rescue of the neural plate is seen (M) compared to triple morphants (L) and uninjected sibling control embryos (K).|
|Figure 5. Ventral Tissues Are Expanded in FCN Morphants(A–P) are lateral views with anterior to the left of neurula (st 13–14) embryos, whereas (Q–F′) are dorsal views with anterior to the top. Expression of multiple ventral markers (BAMBI [A–D and Q–T], msx1 [E–H and U–X], sizzled [I–L and Y–B′], and cytokeratin [M–P and C′–F′]) are shown in uninjected sibling embryos (A, C, E, G, I, K, M, O, Q, S, U, W, Y, A′, C′, and E′) and FCN triple morphants (B, F, J, N, R, V, Z, and D′) as well as in β-catenin morphants (D, H, L, P, T, X, B′, and F′) for comparison.|
|Figure 6. Spemann’s Organizer Is Specified in FCN MorphantsAll panels are vegetal views of early gastrula embryos (st 10.5 [A–T]) or midgastrula (st 11 [U–X]) with dorsal to the right in embryos where dorsal can be distinguished. Expression of multiple mesodermal markers (brachyury [xbra] [A–D], goosecoid [gsc] [E–H], xnr3 [I–L], siamois [M–P], vent2 [Q–T], and myf5 [U–X]) are shown in uninjected sibling embryos (A, C, E, G, I, K, M, O, Q, S, U, and W) and FCN triple morphants (B, F, J, N, R, and V) as well as β-catenin morphants (D, H, L, P, T, and X) for comparison.|
|myod1 (myogenic differentiation 1) is a standard marker for presomitic mesoderm. Note that as somites form in an anterior to posterior direction, myod1 is broadly expressed more posteriorly in this NF stage 14/15 embryo ( dorsal view, anterior up).|