Distinct cranial neural crest-derived cell types (a number of neuronal as well as non-neuronal cell lineages) are generated at characteristic times and positions in the rhombomeres of the hindbrain in developing vertebrate embryos. To examine this developmental process, we developed a novel strategy designed to test the efficacy of gain-of-function Msx2 expression within rhombomeres in ovo prior to the emigration of cranial neural crest cells (CNCC). Previous studies indicate that CNCC from odd-numbered rhombomeres (r3 and r5) undergo apoptosis in response to exogenous BMP4. We provide evidence that targeted infection in ovo using adenovirus containing Msx2 and a reporter molecule indicative of translation can induce apoptosis in either even- or odd-numbered rhombomeres. Furthermore, infected lacZ-control explants indicated that CNCC emigrated, and that 20% of these cells were double positive for crest cell markers HNK-1 and beta-gal. In contrast, there were no HNK-1 and Msx2 double positive cells emigrating from Msx2 infected explants. These results support the hypothesis that apoptotic elimination of CNCC can be induced by ‘gain-of-function’ Msx2 expression in even-numbered rhombomeres. These inductive interactions involve qualitative, quantitative, positional and temporal differences in TGF-beta-related signals, Msx2 expression and other transcriptional control.

Baker
J. C.
,
Harland
R. C.
(
1997
)
From receptor to nucleus: the Smad pathway.
Curr. Opin. Genet. Dev
7
,
467
473
Barlow
A. J.
,
Francis
W. P.
(
1997
)
Ectopic application of recombinant BMP-2 and BMP-4 can change patterning of developing chick facial primordia.
Development
124
,
391
398
Birgbauer
E.
,
Sechrist
J.
,
Bronner
F. M.
,
Fraser
S.
(
1995
)
Rhombomeric origin and rostrocaudal reassortment of neural crest cells revealed by intravital microscopy.
Development
121
,
935
945
Brown
J. M.
,
Robertson
K. E.
,
Wedden
S. E.
,
Tickle
C.
(
1997
)
Alterations in Msx1 and Msx2 expression correlate with inhibition of outgrowth of chick facial promordia induced by retinoic acid.
Anat Embryol
195
,
203
207
Buxton
P.
,
Hunt
P.
,
Ferretti
P.
,
Thorogood
P.
(
1997
)
A role for midline closure in the reestablishment of dorsoventral pattern following dorsal hindbrain ablation.
Dev. Biol
183
,
150
165
Clarke
P. G. H.
,
Clarke
S.
(
1996
)
Nineteenth century research on naturally occurring cell death and related phenomena.
Anat Embryol
193
,
81
99
Couly
G. F.
,
Coltey
P. M.
,
Le
D. N.
(
1993
)
The triple origin of skull in higher vertebrates: a study in quail-chick chimeras.
Development
117
,
409
429
Fanger
G. R.
,
Gerwins
P.
,
Widmann
C.
,
Jarpe
M. B.
,
Johnson
G. L.
(
1997
)
MEKKs, GCKs, MLKs, PAKs, TAKs, and tpls: upstreamregulators of the c-Jun amino-terminal kinases?.
Curr. Opin. Genet. Dev
7
,
67
74
Flenniken
A. M.
,
Gale
N. W.
,
Yancopoulos
G. D.
,
Wilkinson
D. G.
(
1996
)
Distinct and overlapping expression patterns of ligands for Eph-related receptor tyrosine kinases during mouse embryogenesis.
Dev. Biol
179
,
382
401
Foerst-Potts
L.
,
Sadler
T. W.
(
1997
)
Disruption of Msx-1 and Msx-2 reveals roles for these genes in craniofacial, eye, and axial development.
Dev Dyn
209
,
70
84
Gale
N. W.
,
Holland
S. J.
,
Valenzuela
D. M.
,
Flenniken
A.
,
Pan
L.
,
Ryan
T. E.
,
Henkemeyer
M.
,
Strebhardt
K.
,
Hirai
H.
,
Wilkinson
D. G.
,
Pawson
T.
,
Davis
S.
,
Yancopoulos
G. D.
(
1996
)
Eph receptors and ligands comprise two major specificity subclasses and are reciprocally compartmentalized during embryogenesis.
Neuron
17
,
9
19
Ganan
Y.
,
Macias
D.
,
Duterque
C. M.
,
Ros
M. A.
,
Hurle
J. M.
(
1996
)
Role of TGFs and BMPs as signals controlling the position of the digits and the areas of interdigital cell death in the developing chick limb autopod.
Development
122
,
2349
2357
Glozak
M. A.
,
Rogers
M. B.
(
1996
)
Specific induction of apoptosis in P19 embryonal carcinoma cells by retinoic acid and BMP2 or BMP4.
Dev. Biol
179
,
458
70
Graham
A.
,
Heyman
I.
,
Lumsden
A.
(
1993
)
Even-numbered rhombomeres control the apoptotic elimination of neural crest cells from odd-numbered rhombomeres in the chick hindbrain.
Development
119
,
233
245
Graham
A.
,
Francis
W. P.
,
Brickell
P.
,
Lumsden
A.
(
1994
)
The signaling molecule BMP4 mediates apoptosis in the rhombencephalic neural crest.
Nature
372
,
684
686
Graham
A.
,
Koentges
G.
,
Lumsden
A.
(
1996
)
Neural crest apoptosis and the establishment of craniofacial pattern: an honorable death.
Mol. Cell Neurosci
8
,
76
83
Hamburger
V.
,
Hamilton
H. L.
(
1992
)
A series of normal stages in the development of the chick embryo.
Dev. Dyn
195
,
231
72
Hunt
P.
,
Ferretti
P.
,
Krumlauf
R.
,
Thorogood
P.
(
1995
)
Restoration of normal Hox code and branchial arch morphogenesis after extensive deletion of hindbrain neural crest.
Dev. Biol
168
,
584
597
Jacobson
M. D.
,
Weil
M.
,
Raff
M. C.
(
1997
)
Programmed cell death in animal development.
Cell
88
,
347
354
Köntges
G.
,
Lumsden
A.
(
1996
)
Rhombencephalic neural crest segmentation is preserved throughout craniofacial ontogeny.
Development
122
,
3229
3242
Kuratani
S. C.
,
Eichele
G.
(
1993
)
Rhombomere transplantation repatterns the segmental organization of cranial nerves and reveals cell-autonomous expression of a homeodomain protein.
Development
117
,
105
117
Liem
K. F.
,
Tremmi
G.
,
Roelink
H.
,
Jessell
T.
(
1995
)
Dorsal differentiation of neural plate cells induced by BMP-mediated signals from epidermal ectoderm.
Cell
82
,
969
979
Liu
Y. H.
,
Kundu
R.
,
Wu
L.
,
Luo
W.
,
Ignelzi
M. J.
,
Snead
M. L.
,
Maxson
R. J.
(
1995
)
Premature suture closure and ectopic cranial bone in mice expressing Msx2 transgenes in the developing skull.
Proc Natl Acad Sci U S A
92
,
6137
6141
Lumsden
A.
,
Sprawson
N.
,
Graham
A.
(
1991
)
Segmental origin and migration of neural crest cells in the hindbrain region of the chick embryo.
Development
113
,
1281
1291
Macias
D.
,
Ganan
Y.
,
Sampath
T. K.
,
Piedra
M. E.
,
Ros
M. A.
,
Hurle
J. M.
(
1997
)
Role of BMP-2 and OP-1 (BMP-7) in programmed cell death and skeletogenesis during chick limb development.
Development
124
,
1109
1117
Maden
M.
,
Graham
A.
,
Gale
E.
,
Rollinson
C.
,
Zile
M.
(
1997
)
Positional apoptosis during vertebrate CNS development in the absence of endogenous retinoids.
Development
124
,
2799
2805
Marazzi
G.
,
Wang
Y.
,
Sassoon
D.
(
1997
)
Msx2 is a transcriptional regulator in the BMP4-mediated programmed cell death pathway.
Dev. Biol
186
,
127
138
Massague
J.
(
1996
)
TGFsignaling: receptors, transducers, and Mad proteins.
Cell
85
,
947
950
Miyake
S.
,
Makimura
M.
,
Kanegae
Y.
,
Harada
S.
,
Sato
Y.
,
Takamori
K.
,
Tokuda
C.
,
Saito
I.
(
1996
)
Efficient generation of recombinant adenoviruses using adenovirus DNA-terminal protein complex and a cosmid bearing the full-length virus genome.
Proc. Natl. Acad. Sci. USA
93
,
1320
1324
Mizushima
S.
,
Nagata
S.
(
1990
)
pEF-BOS, a powerful mammalian expression vector.
Nucl. Acids Res
18
,
5322
–.
Monsoro
B. A.
,
Duprez
D.
,
Watanabe
Y.
,
Bontoux
M.
,
Vincent
C.
,
Brickell
P.
,
Le
D. N.
(
1996
)
The role of bone morphogenetic proteins in vertebral development.
Development
122
,
3607
3616
Phippard
D. J.
,
Weber
H. S.
,
Sharpe
P. T.
,
Naylor
M. S.
,
Jayatalake
H.
,
Maas
R.
,
Woo
I.
,
Roberts
C. D.
,
Francis
W. P.
,
Liu
Y. H.
,
Maxson
R.
,
Hill
R. E.
,
Dale
T. C.
(
1996
)
Regulation of Msx-1, Msx-2, Bmp-2 and Bmp-4 during foetal and postnatal mammary gland development.
Development
122
,
2729
2737
Reddi
A. H.
(
1997
)
Bone morphogenetic proteins: an unconventional approach to isolation of first mammalian morphogens.
Cytokine Growth Factor Rev
8
,
11
20
Rifas
L.
,
Towler
D. A.
,
Avioli
L. V.
(
1997
)
Gestational exposure to ethanol suppresses msx2 expression in developing mouse embryos.
Proc. Natl. Acad. Sci. USA
94
,
7549
7554
Ros
M. A.
,
Piedra
M. E.
,
Fallon
J. F.
,
Hurle
J. M.
(
1997
)
Morphogenetic potential of the chick leg interdigital mesoderm when diverted from the cell death program.
Dev Dyn
208
,
406
419
Saldivar
J. R.
,
Sechrist
J. W.
,
Krull
C. E.
,
Ruffins
S.
,
Bronner
F. M.
(
1997
)
Dorsal hindbrain ablation results in rerouting of neural crest migration and changes in gene expression, but normal hyoid development.
Development
124
,
2729
2739
Sanders
E. J.
,
Wride
M. A.
(
1995
)
Programmed cell death in development.
Int Rev Cytol
163
,
105
173
Sechrist
J.
,
Serbedzija
G. N.
,
Scherson
T.
,
Fraser
S. E.
,
Bronner
F. M.
(
1993
)
Segmental migration of the hindbrain neural crest does not arise from its segmental generation.
Development
118
,
691
703
Shibuya
H.
,
Yamaguchi
K.
,
Shirakabe
K.
,
Tonegawa
A.
,
Gotoh
Y.
,
Ueno
N.
,
Irie
K.
,
Nishida
E.
,
Matsumoto
K.
(
1996
)
TAB1: an activator of the TAK1 MAPKKK in TGF-signal transduction.
Science
272
,
1179
1182
Sulik
K. K.
,
Cook
C. S.
,
Webster
W. S.
(
1988
)
Teratogens and Craniofacial malformations: relationships to cell death.
Development
103
,
213
231
Tosney
K. W.
(
1982
)
The segregation and early migration of cranial neural crest cells in the avian embryo.
Dev. Biol
89
,
13
24
Vainio
S.
,
Karavanova
I.
,
Jowett
A.
,
Thesleff
I.
(
1993
)
Identification of BMP-4 as a signal mediating secondary induction between epithelial and mesenchymal tissues during early tooth development.
Cell
75
,
45
58
Wang
H. U.
,
Anderson
D. J.
(
1997
)
Eph family transmembrane ligands can mediate repulsive guidance of trunk neural crest migration and motor axon outgrowth.
Neuron
18
,
383
396
Webster
W. S.
,
Johnston
M. C.
,
Lammer
E. J.
,
Sulik
K. K.
(
1986
)
Isotretinoin embryopathy and the cranial neural crest: an in vivo and in vitro study.
J. Craniofac. Genet. Dev. Biol
6
,
211
222
Winograd
J.
,
Reilly
M. P.
,
Roe
R.
,
Lutz
J.
,
Laughner
E.
,
Xu
X.
,
Hu
L.
,
Asakura
T.
,
vander
K. C.
,
Strandberg
J. D.
,
Semenza
G. L.
(
1997
)
Perinatal lethality and multiple craniofacial malformations in MSX2 transgenic mice.
Hum. Mol. Genet
6
,
369
379
Wu
L.
,
Wu
H.
,
Sangiorgi
F.
,
Wu
N.
,
Bell
J. R.
,
Lyons
G. E.
,
Maxon
R.
(
1997
)
Miz1, a novel zinc finger transcription factor that interact with Msx2 and enhance its affinity for DNA.
Mech. Dev
65
,
3
17
Yamaguchi
K.
,
Shirakabe
K.
,
Shibuya
H.
,
Irie
K.
,
Oishi
I.
,
Ueno
N.
,
Taniguchi
T.
,
Nishida
E.
,
Matsumoto
K.
(
1995
)
Identification of a member of the MAPKKK family as a potential mediator of TGF-signal transduction.
Science
270
,
2008
2011
Zhang
H.
,
Hu
G.
,
Wang
H.
,
Sciavolino
P.
,
Iler
N.
,
Shen
M. M.
,
Abate-Shen
C.
(
1997
)
Heterodimerization of Msx and Dlx homeoproteins results in functional antagonism.
Mol. Cell Biol
17
,
2920
2932
Zou
H.
,
Niswander
L.
(
1996
)
Requirement for BMP signaling in interdigital apoptosis and scale formation.
Science
272
,
738
741
Zou
H.
,
Wieser
R.
,
Massague
J.
,
Niswander
L.
(
1997
)
Distinct roles of type I bone morphogenetic protein receptors in the formation and differentiation of cartilage.
Genes Dev
11
,
2191
2203
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