Immediately prior to gastrulation the murine embryo consists of an outer layer of visceral endoderm (VE) and an inner layer of ectoderm. Differentiation and migration of the ectoderm then occurs to produce the three germ layers (ectoderm, embryonic endoderm and mesoderm) from which the fetus is derived. An indication that the VE might have a critical role in this process emerged from studies of Hnf-4(−/−) mouse embryos which fail to undergo normal gastrulation. Since expression of the transcription factor HNF-4 is restricted to the VE during this phase of development, we proposed that HNF-4-regulated gene expression in the VE creates an environment capable of supporting gastrulation. To address this directly we have exploited the versatility of embryonic stem (ES) cells which are amenable to genetic manipulation and can be induced to form VE in vitro. Moreover, embryos derived solely from ES cells can be generated by aggregation with tetraploid morulae. Using Hnf-4(−/−) ES cells we demonstrate that HNF-4 is a key regulator of tissue-specific gene expression in the VE, required for normal expression of secreted factors including alphafetoprotein, apolipoproteins, transthyretin, retinol binding protein, and transferrin. Furthermore, specific complementation of Hnf-4(−/−) embryos with tetraploid-derived Hnf-4(+/+) VE rescues their early developmental arrest, showing conclusively that a functional VE is mandatory for gastrulation.

REFERENCES

Ang
S. L.
,
Rossant
J.
(
1994
)
HNF-3is essential for node and notochord formation in mouse development.
Cell
78
,
561
574
Barton
S. C.
,
Surani
M. A. H.
,
Norris
M. L.
(
1984
)
Role of paternal and maternal genomes in mouse development.
Nature
311
,
374
376
Basheeruddin
K.
,
Stein
P.
,
Strickland
S.
,
Williams
D. L.
(
1987
)
Expression of the murine apolipoprotein E gene is coupled to the differentiated state of F9 embryonal carcinoma cells.
Proc. Natl. Acad. Sci. USA
84
,
709
713
Brent
R. L.
,
Beckman
D. A.
,
Jensen
M.
,
Koszalka
T. R.
(
1990
)
Experimental yolk sac dysfunction as a model for studying nutritional disturbances in the embryo during organogenesis.
Teratology
41
,
405
413
Carmeliet
P.
,
Ferreira
V.
,
Breier
G.
,
Pollefeyt
S.
,
Kieckens
L.
,
Gertsenstein
M.
,
Fahrig
M.
,
Vandenhoeck
A.
,
Harpal
K.
,
Erberhardt
C.
,
Declercq
C.
,
Pawling
J.
,
Moons
L.
,
Collen
D.
,
Risau
W.
,
Nagy
A.
(
1996
)
Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele.
Nature
380
,
435
439
Cereghini
S.
,
Ott
M.-O.
,
Power
S.
,
Maury
M.
(
1992
)
Expression patterns of vHNF1 and HNF1 homeoproteins in early postimplantation embryos suggest distinct and sequential developmental roles.
Development
116
,
783
797
Chen
W. S.
,
Manova
K.
,
Weinstein
D. C.
,
Duncan
S. A.
,
Plump
A. S.
,
Prezioso
V. R.
,
Bachvarova
R. F.
,
Darnell
J. E.
Jr
(
1994
)
Disruption of the HNF-4 gene, expressed in visceral endoderm, leads to cell death in embryonic ectoderm and impaired gastrulation of mouse embryos.
Genes Dev
8
,
2466
2477
Coucouvanis
E.
,
Martin
G. R.
(
1995
)
Signals for death and survival: a two-step mechanism for cavitation in the vertebrate embryo.
Cell
83
,
279
287
Doetschman
T. C.
,
Eistetter
H.
,
Katz
M.
,
Schmidt
W.
,
Kemler
R.
(
1985
)
The in vitro development of blastocyst-derived embryonic stem cell lines: formation of visceral yolk sac, blood islands and myocardium.
J. Embryol. Exp. Morphol
87
,
27
45
Drewes
T.
,
Senkel
S.
,
Holewa
B.
,
Ryffel
G. U.
(
1996
)
Human hepatocyte nuclear factor 4 isoforms are encoded by distinct and differentially expressed genes.
Mol. Cell Biol
16
,
925
931
Duncan
S. A.
,
Manova
K.
,
Chen
W. S.
,
Hoodless
P.
,
Weinstein
D. C.
,
Bachvarova
R. F.
,
Darnell
J. E.
Jr
(
1994
)
Expression of transcription factor HNF-4 in the extraembryonic endoderm, gut, and nephrogenic tissue of the developing mouse embryo: HNF-4 is a marker for primary endoderm in the implanting blastocyst.
Proc. Natl. Acad. Sci. USA
91
,
7598
7602
Episkopou
V.
,
Maeda
S.
,
Nishiguchi
S.
,
Kazunori
S.
,
Gaitanaris
G. A.
,
Gottesman
M. E.
,
Robertson
E. J.
(
1993
)
Disruption of the transthyretin gene results in mice with depressed levels of plasma retinol and thyroid hormone.
Proc. Nat. Acad. Sci. USA
90
,
2375
2379
Evans
M. J.
,
Kaufman
M. H.
(
1981
)
Establishment in culture of pluripotential cells from mouse embryos.
Nature
292
,
154
156
Farese
R. V.
,
Ruland
S. L.
,
Flynn
L. M.
,
Stokowski
R. P.
,
Young
S. G.
(
1995
)
Knockout of the mouse apolipoprotein B gene results in embryonic lethality in homozygotes and protection against diet-induced hypercholesterolemia in heterozygotes.
Proc. Natl. Acad. Sci. USA
92
,
1774
1778
Gardner
R. L.
(
1983
)
Origin and differentiation of extraembryonic endoderm in the mouse.
J. Embryol. Exp. Morphol
80
,
251
288
Grover
A.
,
Andrews
G.
,
Adamson
E. D.
(
1983
)
Role of laminin in epithelium formation by F9 aggregates.
J. Cell Biol
97
,
137
144
Grover
A.
,
Oshima
R. G.
,
Adamson
E. D.
(
1983
)
Epithelial layer formation in differentiating aggregates of F9 embryonal carcinoma cells.
J. Cell Biol
96
,
1690
1696
Guillemot
F.
,
Nagy
A.
,
Auerbach
A.
,
Joyner
A. L.
(
1994
)
Essential role of Mash-2 in extraembryonic development.
Nature
371
,
333
336
Harrison
S. M.
,
Dunwoodie
S. L.
,
Arkell
R. M.
,
Lerach
H.
,
Beddington
R. S. P.
(
1995
)
Isolation of novel tissue-specific genes from cDNA libraries representing the individual constituents of the gastrulating mouse embryo.
Development
121
,
2479
2489
Heath
J. K.
,
Deller
M. J.
(
1983
)
Serum-free culture of PC13 murine embryonal carcinoma cells.
J. Cell. Physiol
115
,
225
230
Hooper
M.
,
Hardy
K.
,
Handyside
A.
,
Hunter
S.
,
Monk
M.
(
1987
)
HPRT-deficient (Lesch-Nyan) mouse embryos derived from germline colonization by cultured cells.
Nature
326
,
292
295
Huang
L. S.
,
Voyiaziakis
E.
,
Markenson
D. F.
,
Sokol
K. A.
,
Hayek
T.
,
Breslow
J. L.
(
1995
)
Apo B gene knockout in mice results in embryonic lethality in homozygotes and neural tube defects, male infertility, and reduced HDL cholesterol ester and Apo A-I transport rates in heterozygotes.
J. Clin. Invest
96
,
2152
2161
Jollie
W. P.
(
1990
)
Development, morphology, and function of the yolk-sac placenta of laboratory rodents.
Terat
41
,
361
381
Kaufman
M. H.
,
Barton
S. C.
,
Surani
A. A. H.
(
1977
)
Normal postimplantation development of mouse parthenogenetic embryos to the forelimb bud stage.
Nature
265
,
53
55
Kubiak
J. C.
,
Tarkowski
A. K.
(
1985
)
Electrofusion of mouse blastomeres.
Exp. Cell. Res
157
,
561
566
Kuo
C. J.
,
Conley
P. B.
,
Chen
L.
,
Sladek
F. M.
,
Darnell
J. E.
Jr.
,
Crabtree
G. R.
(
1992
)
A transcriptional hierarchy involved in mammalian cell-type specification.
Nature
355
,
458
460
Lee
J. J.
,
Radice
G.
,
Perkins
C. P.
,
Constantini
F.
(
1992
)
Identification and characterization of a novel, evolutionarily conserved gene disrupted bythe murine Hβ58 embryonic lethal transgene insertion.
Development
115
,
277
288
Martin
G. R.
(
1981
)
Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells.
Proc. Natl. Acad. Sci. USA
78
,
7634
7638
McGrath
J.
,
Solter
D.
(
1984
)
Completion of mouse embryogenesis requires both the maternal and paternal genomes.
Cell
37
,
179
183
Meehan
R. R.
,
Barlow
D. P.
,
Hill
R. E.
,
Hogan
B. L. M.
,
Hastie
N. D.
(
1984
)
Pattern of serum protein gene expression in the mouse visceral yolk sac and foetal liver.
EMBO J
3
,
1881
1885
Mortensen
R. M.
,
Conner
D. A.
,
Chao
S.
,
Geisterfer-Lowrance
A. A. T.
,
Seidman
J. G.
(
1992
)
Production of homozygous mutant ES cells with a single targeting construct.
Mol. Cell. Biol
12
,
2391
2395
Nagy
A.
,
Gocza
E.
,
Diaz
E. M.
,
Prideaux
V. R.
,
Ivanyi
E.
,
Markkula
M.
,
Rossant
J.
(
1990
)
Embryonic stem cells alone are able to support fetal development in the mouse.
Development
110
,
815
821
Plump
A. S.
,
Smith
j. D.
,
Hayek
T.
,
Aalto-Setaa
K.
,
Walsh
A.
,
Verstuyft
J. G.
,
Rubin
E. M.
,
Breslow
J. L.
(
1992
)
Severe hypercholesterolemia and atherosclerosis in apolipoprotein E-deficient mice created by homologous recombination in ES cells.
Cell
71
,
343
353
Pontoglio
M.
,
Barra
J.
,
Hadchouei
M.
,
Doyen
A.
,
Kress
C.
,
Bach
J. P.
,
Babinet
C.
,
Yaniv
M.
(
1996
)
Hepatocyte nuclear factor 1 inactivation results in hepatic dysfunction, phenylketonuria and renal Fanconi syndrome.
Cell
84
,
575
585
Sato
M.
,
Muramatsu
T.
(
1985
)
Reactivity of five N-acetylgalactosamine-recognizing lectins with preimplantation embryos, early postimplantation embryos, and teratocarcinoma cells of the mouse.
Diff
29
,
29
38
Sladek
F. M.
,
Zhong
W.
,
Lai
E.
,
Darnell
J. E.
Jr
(
1990
)
Liver-enriched transcription factor HNF-4 is a novel member of the steroid hormone receptor superfamily.
Genes Dev
4
,
2353
2365
Soudais
C.
,
Bielinska
M.
,
Heikinheimo
M.
,
MacArthur
C. A.
,
Narita
N.
,
Saffitz
J. E.
,
Simon
M. C.
,
Leiden
J. M.
,
Wilson
D. B.
(
1995
)
Targeted mutagenesis of the transcription factor GATA-4 gene in mouse embryonic stem cells disrupts visceral endoderm differentiation in vitro.
Development
121
,
3877
3888
Spyropoulos
D. D.
,
Capecchi
M. R.
(
1994
)
Targeted disruption of the even-skipped gene, evx1, causes early postimplantation lethality of the mouse conceptus.
Genes Dev
8
,
1949
1961
Sturm
K.
,
Flannery
M. L.
,
Pederson
R. A.
(
1994
)
Abnormal development of embryonic and extraembryonic cell lineages in parthenogenetic mouse embryos.
Dev. Dyn
201
,
11
28
Surani
M. A. H.
,
Barton
S. C.
,
Norris
M. L.
(
1984
)
Development of reconstituted mouse eggs suggests imprinting of the genome during gametogenesis.
Nature
308
,
548
308
Taraviras
S.
,
Monaghan
A. P.
,
Schutz
G.
,
Kelsey
G.
(
1994
)
Characterization of the mouse HNF-4 gene and its expression during mouse embryogenesis.
Mech. Dev
48
,
67
79
Tian
J. M.
,
Schibler
U.
(
1991
)
Tissue-specific expression of the gene encoding hepatocyte nuclear factor 1 may involve hepatocyte nuclear factor 4.
Genes Dev
5
,
2225
2234
Weinstein
D. C.
,
Altaba
A. R. i.
,
Chen
W. S.
,
Hoodless
P.
,
Prezioso
V. R.
,
Jessell
T. M.
,
Darnell
J. E.
Jr
(
1994
)
The winged-helix transcription factor HNF-3is required for notochord development in the mouse embryo.
Cell
78
,
575
588
Williamson
R.
,
Lee
D.
,
Hagaman
J.
,
Maeda
N.
(
1992
)
Marked reduction in high density lipoprotein cholesterol in mice genetically modified to lack apolipoprotein A-I.
Proc. Natl. Acad. Sci. USA
89
,
7134
7138
Wilson
P. A.
,
Melton
D. A.
(
1994
)
Mesodermal patterning by an inducer gradient depends on secondary cell-cell communication.
Curr. Biol
4
,
676
686
Wu
T.-C.
,
Wan
Y.-J.
,
Damjanov
I.
(
1983
)
Flourescein-conjugated Badeira simplicifolia lectin as a marker of endodermal, yolk sac and trophoblastic differentiaition in the mouse embryo.
Diff
24
,
55
59
Zhong
W.
,
Sladek
F. M.
,
Darnell
J. E.
Jr
(
1993
)
The expression pattern of a Drosophila homolog to the mouse transcription factor HNF-4 suggests a determinative role in gut formation.
EMBO J
12
,
537
544
This content is only available via PDF.