The dorsal gastrula organizer plays a fundamental role in establishment of the vertebrate axis. We demonstrate that the zebrafish bozozok (boz) locus is required at the blastula stages for formation of the embryonic shield, the equivalent of the gastrula organizer and expression of multiple organizer-specific genes. Furthermore, boz is essential for specification of dorsoanterior embryonic structures, including notochord, prechordal mesendoderm, floor plate and forebrain. We report that boz mutations disrupt the homeobox gene dharma. Overexpression of boz in the extraembryonic yolk syncytial layer of boz mutant embryos is sufficient for normal development of the overlying blastoderm, revealing an involvement of extraembryonic structures in anterior patterning in fish similarly to murine embryos. Epistatic analyses indicate that boz acts downstream of beta-catenin and upstream to TGF-beta signaling or in a parallel pathway. These studies provide genetic evidence for an essential function of a homeodomain protein in beta-catenin-mediated induction of the dorsal gastrula organizer and place boz at the top of a hierarchy of zygotic genes specifying the dorsal midline of a vertebrate embryo.

Reference

Acampora
D.
,
Mazan
S.
,
Lallemand
Y.
,
Avantaggiato
V.
,
Maury
M.
,
Simeone
A.
,
Prulet
P.
(
1995
)
Forebrain and midbrain regions are deleted in Otx/ mutants due to a defective anterior neuroectoderm specification during gastrulation.
Development
121
,
3279
3290
Ang
S.-L.
,
Jin
O.
,
Rhinn
M.
,
Daigle
N.
,
Stevenson
L.
,
Rossant
J.
(
1996
)
A targeted Otx2 mutation leads to severe defects in gastrulation and formation of axial mesoderm and to deletion of rostral brain.
Development
122
,
243
252
Beddington
R. S. P.
,
Robertson
E. J.
(
1998
)
Anterior patterning in the mouse.
Trends in Genetics
14
,
277
284
Blagden
C. S.
,
Currie
P. D.
,
Ingham
P. W.
,
Hughes
S. M.
(
1997
)
Notochord induction of zebrafish slow muscle mediated by Sonic hedgehog.
Genes Dev
11
,
2163
2175
Bouwmeester
T.
,
Leyns
L.
(
1997
)
Vertebrate head induction by anterior primitive endoderm.
BioEssays
19
,
855
863
Brannon
M.
,
Kimelmann
D.
(
1996
)
Activation of Siamois by the Wnt Pathway.
Dev. Biol
180
,
344
347
Cooper
M. S.
,
D'Amico
L. A.
(
1996
)
A cluster of noninvoluting endocytic cells at the margin of the zebrafish blastoderm marks the site of embryonic shield formation.
Dev. Biol
180
,
184
198
Crease
D. J.
,
Dyson
S.
,
Gurdon
J. B.
(
1998
)
Cooperation between the activin and Wnt pathways in the spatial control of organizer gene expression.
Proc. Natl Acad. Sci. USA
95
,
4398
4403
Dale
L.
,
Slack
J. M. W.
(
1987
)
Regional specification within the mesoderm of early embryos of Xenopus laevis.
Development
100
,
279
295
Dohrmann
C. E.
,
Kessler
D. S.
,
Melton
D. A.
(
1996
)
Induction of axial mesoderm by zDVR-1, the zebrafish orthologue of Xenopus Vg-1.
Dev. Biol
175
,
108
117
Erter
C. E.
,
Solnica-Krezel
L.
,
Wright
C. V. E.
(
1998
)
Zebrafish nodal-related 2 encodes an early mesendodermal inducer signaling from the extraembryonic YSL.
Dev. Biol
204
,
361
372
Fan
M. J.
,
Sokol
S. Y.
(
1997
)
A role for Siamois in Spemann organizer formation.
Development
124
,
2581
1589
Feldman
B.
,
Gates
M. A.
,
Egan
E. S.
,
Dougan
S. T.
,
Rennebeck
G.
,
Sirotkin
H. I.
,
Schier
A. F.
,
Talbot
W. S.
(
1998
)
Zebrafish organizer development and germ-layer formation require nodal-related signals.
Nature
395
,
181
185
Gumbiner
B. M.
(
1995
)
Signal transduction by b-catenin.
Current Biol
7
,
634
640
Hammerschmidt
M.
,
Bitgood
M. J.
,
McMahon
A. P.
(
1996
)
Protein kinase A is a common negative regulator of hedgehog signaling in the vertebrate embryo.
Genes Dev
10
,
647
658
Harland
R.
,
Gerhart
J.
(
1997
)
Formation and function of Spemann's organizer.
Annu Rev. Cell Dev. Biol
13
,
611
667
Heasman
J.
(
1997
)
Patterning in Xenopus blastula.
Development
124
,
4179
4191
Heasman
J.
,
Crawfor
A.
,
Goldstone
K.
,
Garner-Hamrick
P.
,
Gumbiner
B.
,
McCrea
P.
,
Kintner
C.
,
Noro
C. Y.
,
Wylie
C.
(
1994
)
Overexpression of cadherins and underexpression of-catenin inhibit dorsal mesoderm induction in early Xenopus embryos.
Cell
79
,
791
803
Hedgepeth
C. M.
,
Conrad
L. J.
,
Zhang
J.
,
Huang
H. C.
,
Lee
V. M.
,
Klein
P. S.
(
1997
)
Activation of the Wnt signaling pathway: a molecular mechanism for lithium action.
Dev. Biol
185
,
82
91
Jesuthasan
S.
,
Strahle
U.
(
1997
)
Dynamic microtubules and specification of the zebrafish embryonic axis.
Curr. Biol
7
,
31
42
Kelly
G. M.
,
Erezyilmaz
D. F.
,
Moon
R. T.
(
1995
)
Induction of a secondary embryonic axis in zebrafish occurs following the overexpression of-catenin.
Mech. Dev
53
,
261
273
Kessler
D. D.
,
Melton
D. A.
(
1995
)
Induction of dorsal mesoderm by soluble, mature Vg1 protein.
Development
121
,
2155
2164
Kessler
D. S.
(
1997
)
Siamois is required for formation of Spemann's organizer.
Proc. Natl Acad. Sci. USA
94
,
13017
13022
Kimelman
D.
,
Christian
J. L.
,
Moon
R. T.
(
1992
)
Synergistic principles of development: overlapping patterning systems in Xenopus mesoderm induction.
Development
116
,
1
9
Kimmel
C. B.
,
Ballard
W. W.
,
Kimmel
S. R.
,
Ullmann
B.
,
Schilling
T. F.
(
1995
)
Stages of embryonic development of the zebrafish.
Dev. Dynamics
203
,
253
310
Knapik
E.
,
Goodman
A.
,
Atkinson
O. S.
,
Roberts
C. T.
,
Shiozawa
M.
,
Sim
C. U.
,
Weksler-Zangen
S.
,
Trolliet
M. R.
,
Futrell
C.
,
Innes
B. A.
,
Koike
G.
,
McLaughlin
M. G.
,
Pierre
L.
,
Simon
J. S.
,
Vilalonga
E.
,
Roy
M.
,
Chiang
P.-W.
,
Fishman
M. C.
,
Driever
W.
,
Jacob
H. J.
(
1996
)
A reference cross for zebrafish (Danio rerio) anchored with simple sequence length polymorphisms.
Development
123
,
451
460
Knapik
E. W.
,
Goodman
A.
,
Ekker
M.
,
Chevrette
M.
,
Delgado
J.
,
Neuhauss
S.
,
Shimoda
N.
,
Driever
W.
,
Fishman
M. C.
,
Jacob
H.
(
1998
)
A microsatellite genetic linkage map for zebrafish (Danio rerio).
Nature Genetics
18
,
338
343
Krauss
S.
,
Concordet
J. P.
,
Ingham
P. W.
(
1993
)
A functionally conserved homolog of the Drosophila segment polarity gene hh is expressed in tissues with polarizing activity in zebrafish embryos.
Cell
75
,
1431
1444
Larabell
C. A.
,
Torres
M.
,
Rowning
B. A.
,
Yost
C.
,
Miller
J. R.
,
Wu
M.
,
Kimelman
D.
,
Moon
R. T.
(
1997
)
Establishment of the dorso-ventral axis in Xenopus embryos is presaged by early asymmetries in beta-catenin that are modulated by the Wnt signaling pathway.
J. Cell Biol
136
,
1123
1136
Laurent
M. N.
,
Blitz
I. L.
,
Hashimoto
C.
,
Rothbacher
U.
,
Cho
K. W.
(
1997
)
The Xenopus homeobox gene twin mediates Wnt induction of goosecoid in establishment of Spemann's organizer.
Development
124
,
4905
4916
Lemaire
P.
,
Garret
N.
,
Gurdon
J. B.
(
1995
)
Expression cloning of Siamois, a Xenoups homobox gene expressed in dorsal-vegetal cells of blastulae and able to induce a complete secondary axis.
Cell
81
,
85
94
Lemaire
P.
,
Kodjabachian
L.
(
1996
)
The vertebrate organizer: structure and molecules.
Trends Genet
12
,
525
531
Marlow
F.
,
Zwartkruis
F.
,
Malicki
J.
,
Neuhauss
S. C. F.
,
Abbas
L.
,
Weaver
M.
,
Driever
W.
,
Solnica-Krezel
L.
(
1998
)
Functional Interactions of genes mediating convergent extension, knypek and trilobite, during partitioning of the eye primordium in zebrafish.
Dev. Biol
203
,
382
399
Matsuo
I.
,
Kuratani
S.
,
Kimura
C.
,
Takeda
N.
,
Aizawa
S.
(
1995
)
Mouse Otx2 functions in the formation and patterning of rostral head.
Genes Dev
9
,
2646
2658
Mizuno
T.
,
Yamaha
E.
,
Wakahara
M.
,
Kuroiwa
A.
,
Takeda
H.
(
1996
)
Mesoderm induction in zebrafish.
Nature
383
,
131
132
Moon
R. T.
,
Kimelman
D.
(
1998
)
From cortical rotation to organizer gene expression: toward a molecular explanation of axis specification in Xenopus.
BioEssays
20
,
536
545
Neave
B.
,
Holder
N.
,
Patient
R.
(
1997
)
A graded response to BMP-4 spatially coordinates patterning of the mesoderm and ectoderm in the zebrafish.
Mechan. Dev
62
,
183
195
Nguyen
V. H.
,
Schmid
B.
,
Trout
J.
,
Connors
S. A.
,
Ekker
M.
,
Mullins
M. C.
(
1998
)
Ventral and lateral regions of the zebrafish gastrula, including the neural crest progenitors, are established by a bmp2b/swirl pathway of genes.
Dev. Biol
199
,
93
110
Nieuwkoop
P. D.
(
1973
)
The organization center of the amphibian embryo: its origin, spatial organization, and morphogenetic action.
Adv. Morph
10
,
1
39
Oxtoby
E.
,
Jowett
T.
(
1993
)
Cloning of the zebrafish krox-20 gene (krx-20) and its expression during hindbrain development.
Nucleic Acids Research
21
,
1087
1095
Postlethwait
J. H.
,
Johnson
S. L.
,
Midson
C. N.
,
Talbot
W. S.
,
Gates
M.
,
Ballinger
E. W.
,
Africa
D.
,
Andrews
R.
,
Carl
T.
,
Eisen
J. S.
,
Horne
S.
,
Kimmel
C. B.
,
Hutchinson
M.
,
Johnson
M.
,
Rodriguez
A.
(
1994
)
A genetic linkage map for the zebrafish.
Science
264
,
699
703
Rebagliati
M. R.
,
Toyama
R.
,
Fricke
C.
,
Haffter
P.
,
Dawid
I. B.
(
1998
)
Zebrafish nodal-related genes are implicated in axial patterning and establishing left-right asymmetry.
Dev. Biol
199
,
261
272
Rebagliati
M. R.
,
Toyama
R.
,
Haffter
P.
,
Dawid
I. B.
(
1998
)
cyclops encodes a nodal-related factor involved in midline signaling.
Proc. Natl. Acad. Sci. USA
95
,
9932
9937
Renucci
A.
,
Lemarchandel
V.
,
Rosa
F.
(
1996
)
An activated form of type I serine/threonine kinase receptor TARAM-A reveals a specific signalling pathway involved in fish head organizer formation.
Development
122
,
3735
3743
Rowning
B. A.
,
Wells
J.
,
Wu
M.
,
Gerhard
J. C.
,
Moon
R. T.
,
Larabell
C. A.
(
1997
)
Microtubule-mediated transport of organells and localization of b-catenin to the future dorsal side of Xenopus eggs.
Proc. Natl. Acad. Sci. USA
94
,
1224
1229
Sampath
K.
,
Rubinstein
A. L.
,
Cheng
A. M. S.
,
Liang
J. O.
,
Fekany
K.
,
Solnica-Krezel
L.
,
Korzh
V.
,
Halpern
M. E.
,
Wright
C. V. E.
(
1998
)
Induction of the zebrafish ventral brain and floor plate requires Cyclops/Nodal Signaling.
Nature
395
,
185
189
Schier
A. F.
,
Talbot
W. S.
(
1998
)
The Zebrafish Organizer.
Curr. Op. in. Gen. Dev
8
,
464
471
Schneider
S.
,
Steinbesser
H.
,
Warga
R. M.
,
Hausen
P.
(
1996
)
-catenin translocation into nuclei demarcates the dorsalizing centers in frog and fish embryos.
Mechan. Dev
57
,
191
198
Schulte-Merker
S.
,
Hammerschmidt
M.
,
Beuchle
D.
,
Cho
K. W.
,
Derobertis
E. M.
,
Nusslein-Volhard
C.
(
1994
)
Expression of zebrafish goosecoid and no tail gene products in wild-type and mutant no tail embryos.
Development
120
,
843
852
Schulte-Merker
S.
,
Ho
R. K.
,
Herrmann
B. G.
,
Nusslein-Volhard
C.
(
1992
)
The protein product of the zebrafish homologue of the mouse T gene is expressed in nuclei of the germ ring and the notochord of the early embryo.
Development
116
,
1021
1032
Schulte-Merker
S.
,
Lee
K. J.
,
McMahon
A. P.
,
Hammerschmidt
M.
(
1997
)
The zebrafish organizer requires chordino.
Nature
387
,
862
863
Shawlot
W.
,
Behringer
R.R.
(
1995
)
Requirement for Lim1 in head-organizer function.
Nature
374
,
425
430
Shih
J.
,
Fraser
S. E.
(
1996
)
Characterizing the zebrafish organizer: microsurgical analysis at the early-shield stage.
Development
122
,
1313
1322
Smith
J. C.
,
Price
B. M. J.
,
Van Nimmen
K.
,
Huylebroeck
D.
(
1990
)
Identification of a potent Xenopus mesoderm-inducing factor as a homologue of activin A.
Nature
345
,
729
731
Solnica-Krezel
L.
,
Driever
W.
(
1994
)
Microtubule arrays of the zebrafish yolk cell: organization and function during epiboly.
Development
120
,
2443
2455
Solnica-Krezel
L.
,
Schier
A. F.
,
Driever
W.
(
1994
)
Efficient recovery of ENU-induced mutations from the zebrafish germline.
Genetics
136
,
1401
1420
Solnica-Krezel
L.
,
Stemple
D. L.
,
Mountcastle-Shah
E.
,
Rangini
Z.
,
Neuhauss
S. C. F.
,
Malicki
J.
,
Schier
A.
,
Stainier
D. Y. R.
,
Zwartkruis
F.
,
Abdelilah
S.
,
Driever
W.
(
1996
)
Mutations affecting cell fates and cellular rearrangements during gastrulation in zebrafish.
Development
123
,
117
128
Stachel
S. E.
,
Grunwald
D. J.
,
Myers
P. Z.
(
1993
)
Lithium perturbation and goosecoid expression identify a dorsal specification pathway in the pregastrula zebrafish.
Development
117
,
1261
1274
Strahle
U.
,
Blader
P.
,
Ingham
P. W.
(
1996
)
Expression of axial and sonic hedgehog in wildtype and midline defective zebrafish embryos.
Int. J. Dev. Biol
40
,
929
940
Talbot
W. S.
,
Trevarrow
W.
,
Halpern
M. E.
,
Melby
A. E.
,
Farr
G.
,
Postlethwait
J. H.
,
Jowett
T.
,
Kimmel
C. B.
,
Kimelman
D.
(
1995
)
A homeobox gene essential for zebrafish notochord development.
Nature
378
,
150
157
Thisse
C.
,
Thisse
B.
,
Halpern
M. E.
,
Postlethwait
J. H.
(
1994
)
goosecoid expression in neuroectoderm and mesendoderm is disrupted in zebrafish cyclops gastrulas.
Dev. Biol
164
,
420
429
Thomas
P.
,
Beddington
R.
(
1996
)
Anterior primitive endoderm may be responsible for patterning the anterior neural plate in the mouse embryo.
Current Biology
6
,
1487
1496
Thomsen
G. H.
(
1997
)
Antagonism within and around the organizer: BMP inhibitors in vertebrate body patterning.
Trends in Genetics
13
,
209
211
Toyama
R.
,
Oconnell
M. L.
,
Wright
C. V. E.
,
Kuehn
M. R.
,
Dawid
I. B.
(
1995
)
Nodal induces ectopic goosecoid and lim1 expression and axis duplication in zebrafish.
Development
121
,
383
391
Varlet
I.
,
Collignon
J.
,
Robertson
E. J.
(
1997
)
nodal expression in the primitive endoderm is required for specification of the anterior axis during mouse gastrulation.
Development
124
,
1033
1044
Waldrip
W. R.
,
Bikoff
E. K.
,
Hoodless
P. A.
,
Wrana
J. L.
,
Robertson
E. J.
(
1998
)
Smad2 signaling in extraembryonic tissues determines anterior-posterior polarity of the early mouse embryo.
Cell
92
,
797
808
Watabe
T.
,
Kim
S.
,
Candia
A.
,
Rothbacher
U.
,
Hashimoto
C.
,
Inue
K.
,
Cho
K. W. Y.
(
1995
)
Molecular mechanisms of Spemann's organizer formation: conserved growth factor synergy between Xenopus and mouse.
Genes Dev
9
,
3038
3050
Wylie
C.
,
Kofron
M.
,
Payne
C.
,
Anderson
R.
,
Hosobuchi
M.
,
Joseph
E.
,
Heasman
J.
(
1996
)
Maternal-catenin establishes a ‘dorsal signal’ in early Xenopus embryos.
Development
122
,
2987
2996
Xu
Q.
,
Holder
N.
,
Patient
R.
,
Wilson
S. W.
(
1994
)
Spatially regulated expression of three receptor tyrosine kinase genes during gastrulation in the zebrafish.
Development
120
,
287
299
Yamanaka
Y.
,
Mizuna
T.
,
Sasai
Y.
,
Khishi
M.
,
Takeda
H.
,
Kim
C.-H.
,
Hibi
M.
,
Hirano
T.
(
1998
)
A novel homeobox gene, dharma, can induce the organizer in a non-cell-autonomous manner.
Genes Dev
12
,
2345
2353
Yan
Y.-L.
,
Hatta
K.
,
Riggleman
B.
,
Postlethwait
J. H.
(
1995
)
Expression of a type II collagen gene in the zebrafish embryonic axis.
Dev. Dynamics
203
,
363
376
This content is only available via PDF.