1 RIKEN Center for Developmental Biology, 2-2-3 Minatojima Minamimachi, Chuo-ku,
Kobe 650-0047, Japan
2 Department of Cell and Developmental Biology, Graduate School of Biostudies,
Kyoto University, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, Japan
Author for correspondence (e-mail:
nakagawas{at}riken.jp)
Accepted 7 April 2005
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SUMMARY |
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Key words: Wnt, retina, progenitor cells, stem cells, proneural gene, Notch, Delta, differentiation, cell cycle
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Introduction |
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Wnt family proteins are secreted signaling molecules that regulate various
aspects of developmental processes (reviewed by
Wodarz and Nusse, 1998). Wnt2b
is expressed in the anterior rim of the optic vesicles, neighboring the
marginal progenitor cells of the retina
(Zakin et al., 1998
;
Jasoni et al., 1999
). In mice,
the canonical Wnt pathway that induces target gene transcription (reviewed by
Wodarz and Nusse, 1998
) is
activated in the peripheral part of the retina, as was revealed by expression
of a reporter gene under the control of Wnt-responsive promoter sequences
(Liu et al., 2003
). In chicken
embryos, expression of Lef1 mRNA is upregulated when the canonical Wnt pathway
is activated (Schmidt et al.,
2000
), and this tentative marker for the active state of Wnt
signaling is highly expressed in the marginal, but not in the central, region
of the neural retina (Kubo et al.,
2003
). Such observations suggest that the canonical Wnt signaling
is operating in the marginal part of the optic vesicles where undifferentiated
retinal progenitor cells reside. Wnt2b can inhibit the differentiation of the
progenitor cells derived from the central retina both in vitro and in ovo
(Nakagawa et al., 2003
;
Kubo et al., 2003
). In
addition, blocking of the Wnt signaling pathway by a dominant-negative form of
Lef1, a downstream effector of the canonical Wnt signaling, induces premature
neuronal differentiation in the marginal retina
(Kubo et al., 2003
).
Furthermore, Wnt2b promotes prolonged proliferation of multipotent progenitor
cells in clonal cultures (Kubo et al.,
2003
). These observations indicate that Wnt2b is responsible for
the maintenance of progenitor cells in the marginal retina; however, the
precise molecular mechanism by which Wnt2b maintains progenitor cells in the
undifferentiated state has remained an open question.
A number of proneural genes that encode basic helix-loop-helix (bHLH)
transcription factors are expressed in the developing retina, and they control
the differentiation of particular cell types in combination with other
regulatory factors (reviewed by Vetter and
Brown, 2001; Perron and
Harris, 2000a
; Kageyama et
al., 1997
). For example, forced expression of an atonal family
transcription factor, Ath5, leads to an overproduction of retinal ganglion
cells in various vertebrate species (Liu
et al., 2001
; Matter-Sadzinski
et al., 2001
; Kanekar et al.,
1997
), and this cell type is absent in Ath5-mutant
animals (Kay et al., 2001
;
Brown et al., 2001
;
Wang et al., 2001
). Other
proneural genes that influence the production of specific neuronal cell types
include NeuroD (Yan and Wang,
1998
; Morrow et al.,
1999
; Hutcheson and Vetter,
2001
; Inoue et al.,
2002
), Ath3 (Perron
et al., 1999
; Tomita et al.,
2000
; Inoue et al.,
2002
), neurogenin 2 (Ngn2)
(Marquardt et al., 2001
) and
an acute-scute family transcription factor Ash1
(Tomita et al., 1996
;
Brown et al., 1998
;
Tomita et al., 2000
). The
expression or activities of these proneural genes are regulated by various
mechanisms depending on each gene or the developmental stage. The signal
mediated by the Delta ligand and Notch receptor is an important factor that
antagonizes the proneural gene function through a downstream effector gene
called hairy-related proteins (Hes) (reviewed by
Kageyama et al., 2000
;
Vetter and Brown, 2001
;
Perron and Harris, 2000a
). In
the vertebrate retina, downregulation of Notch expression by antisense
oligonucleotide treatment increases the number of retinal ganglion cells
(Austin et al., 1995
), whereas
persistent activation of Notch signaling by a constitutively active form of
the receptor inhibits neuronal differentiation
(Dorsky et al., 1995
;
Bao and Cepko, 1997
;
Austin et al., 1995
). In
addition, overexpression of Delta inhibits differentiation of retinal
progenitor cells, and that of the dominant-negative form of the ligand leads
to premature neuronal differentiation
(Henrique et al., 1997
;
Dorsky et al., 1997
;
Ahmad et al., 1997
). All of
these experiments support the idea that the Delta-Notch signaling keeps
retinal progenitor cells in an undifferentiated state by antagonizing the
proneural gene cascade and that they differentiate into specific neurons once
they escape this inhibitory signal.
In this study, we first compared the effect of Wnt2b and Delta on retinal progenitor cell proliferation and differentiation. We show that the overexpression of Wnt2b supported prolonged proliferation of retinal progenitor cells whereas that of Delta did not. We also demonstrate that Wnt2b inhibited neuronal differentiation even under the conditions where the Delta/Notch signaling pathway was blocked. Wnt2b downregulated mRNA expression of multiple proneural genes as well as that of Notch1. In addition, forced expression of Cath5 under the control of ubiquitous promoter neutralized the neural differentiation-inhibiting effect of Wnt2b. We propose that Wnt2b maintains progenitor cells by preventing them from entering the differentiation cascade regulated by Notch and proneural genes.
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Materials and methods |
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Explant culture of neural retina
The culture medium used was a 1:1 mixture of Dulbecco's Modified Eagle's
medium and Ham F12 (Nissui, Japan) supplemented with 10% fetal calf serum and
penicillin/streptomycin (DH10). Neural retina of 1 mm2 was taken
from the equatorial region temporal to the optic stalk, and further cut into
pieces of 250 µm2 surface area with fine scissors. In most
cases, some region of the Wnt2b-expressing neural retina formed folds. The
tissue explants were prepared from unfolded regions in that case. The retinal
explants were then placed in the wells of non-coated 24-well dishes (Iwaki,
Japan) with 1 ml of DH10, and the dishes were rotated at a speed of 60 r.p.m.
on a rotary shaker. Half of the culture medium was changed every day. In some
experiments, recombinant EGF and bFGF (Gibco) were added at a concentration of
20 ng/ml each. For labeling proliferating cells, BrdU (Sigma) was added to the
medium at a concentration of 2 µM, and the retinal explants were then
incubated for 30 minutes at 37°C. To block the signaling pathway
downstream of Notch, we used a -secretase inhibitor DAPT
{N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine-t-butyl ester;
Calbiochem}, which was diluted in DMSO and added at a concentration of 2
µM. To obtain Frizzled 8-CRD, we transfected COS7 cells with pJCH100, which
encodes mouse frizzled 8 CRD domain fused to human immunoglobulin heavy chain
(Hsieh et al., 1999
). The
fusion protein was purified on a protein A column, and added to the retinal
cultures at a concentration of 50 µg/ml.
Immunohistochemistry and in situ hybridization
The antibodies used were as follows: mouse anti-Hu (clone 16A11; Molecular
Probe); mouse anti-middle-molecular-weight neurofilament (clone RMO270;
Zymed); mouse anti-Syntaxin (HPC1; Sigma); mouse anti-Glutamine Synthetase
(clone 6; Transduction Laboratory); mouse anti-Pax6 (DSHB); mouse anti-islet1
(clone 40.2D6; DSHB); mouse monoclonal antibody 115A10 (a kind gift from Dr
Fujita); mouse anti-BrdU (clone BU33; Sigma); mouse anti-radial glia-specific
antibody (clone 3CB2; DSHB); mouse anti-Vimentin (clone H5, DSHB), sheep
anti-Chx10 (Exalpha); rabbit anti-Visinin (a kind gift from Dr Miki); rabbit
anti-phosphorylated Histone H3 (Upstate Biology); Cy3-conjugated anti-rabbit
IgG (Chemicon); Alexa 488-conjugated anti-sheep IgG (Molecular Probes) and
Alexa 488-conjugated anti-mouse IgG (Molecular Probes).
The tissue explants or embryos were fixed for 1 hour at room temperature in 4% paraformaldehyde in PBS, permeabilized in 100% methanol for 5 minutes at 20°C, and processed using a standard immunostaining protocol.
For BrdU staining, the sections were treated with 2 N HCl for 2 hours at room temperature. Confocal images were collected using a LSM510 microscope (Zeiss).
For in situ hybridization studies, embryos were fixed in 4% paraformaldehyde overnight at 4°C and processed using standard protocols. The cRNA probes for cNotch1 were kindly provided by Dr. Wakamatsu (Tohoku University, Japan). For Cath5 and NeuroM, the cDNA fragments corresponding to the coding sequences of the proteins were amplified by PCR and subcloned into pCRII (Invitrogen) to make the probes.
RT-PCR analysis of proneural gene expression
Total RNAs were extracted using RNAzol (Tel-Test, Inc., Friendswood, TX,
USA) from E5.5 retinas that had been electroporated with either RCASBP(A)
Wnt2b or control RCASBP(A) provirus. cDNAs were synthesized from 1 µg of
total RNAs using oligo(dT) primer and RevetraAce (TOYOBO) according to the
manufacturer's instructions. One-nanogram of the resultant first-strand cDNAs
were subsequently used for PCR analysis with the following primer pairs:
Notch1, 5'-tggagccactgacgttttgt-3' and
5'-agggatgtcaccataaccat-3'; Delta1,
5'-gaacagccagtttccctgga-3' and
5'-gacgtacaccgactggtact-3'; Cath5,
5'-cgtgctggtgtcttacacta-3' and
5'-ttcacacaagcgtctcattc-3'; NeuroM,
5'-gaccttattcctggagaagc-3' and
5'-cgcttcgctactcgttgaag-3'; Cash1,
5'-gtatcctccctgaccagttt-3' and
5'-cgctcttctgcgtttggaca-3'; Neurogenin1,
5'-ccatacgacttggaaaaccc-3' and
5'-agtggctcaataccccgtgt-3'; Neurogenin2,
5'-aggggtccaggttagaagtc-3' and
5'-ccctttgctcttcagccgta-3'; Lef1,
5'-ctcacacaactggaatccct-3' and
5'-tggtgtcgccgcctttgctt-3'; Gapdh,
5'-acatcatcccagcgtccact-3' and
5'-gatgtaaggtggtacaca-3'. PCR was carried out under cycle
conditions of 94°C for 30 seconds, 50°C 30 seconds, and 72°C for 3
minutes. The number of the cycles was carefully determined for each gene so
that the resultant products were amplified within a linear range: 25 cycles
for Notch1, Delta1, Cath5, Cash1 and Gapdh; 30 cycles for
NeuroM, Ngn2, and Lef1; and 35 cycles for Ngn1. The
sequences of the PCR products were checked by direct sequencing using each of
the primers. For semi-quantitative analysis, the PCR products were run on a 2%
agarose gel, and intensities of corresponding bands were measured by
ImageMaster TotalLab (Amersham biosciences) after staining with ethidium
bromide.
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Results |
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Wnt2b inhibits progenitor cell differentiation in the absence of Delta-Notch signaling
The observations so far mentioned suggest that Wnt2b inhibited
differentiation of retinal progenitor cells through a pathway independent of
the inhibitory signals mediated by Delta/Notch. If that was truly the case, it
can be predicted that Wnt2b might inhibit progenitor cell differentiation in
the absence of the Delta/Notch signaling pathway. To address this issue, we
prepared retinal explants from earlier embryos (E5) and treated them for 24
hours with the -secretase inhibitor DAPT, which prevents the cleavage
of the intracellular portion of Notch and thus functions as a Notch-signaling
inhibitor (Dovey et al., 2001
;
Geling et al., 2002
). In the
untreated control explants, many cells co-expressed Pax6 and Chx10 and
incorporated BrdU (Fig.
3E,I,U), suggesting that the E5 retinal explants had not fully
differentiated yet under this culture condition. In the DAPT-treated control
explants, characteristic epithelial structures of neural retinas were lost,
and spherical aggregates developed that were composed of differentiated
neurons (Fig. 3B,F). In the
aggregates, almost all the cells expressed either visinin or Hu, and few cells
co-expressed the progenitor cell marker Pax6 and Chx10
(Fig. 3B,F). In addition,
BrdU-positive cells were rarely observed
(Fig. 3J,U), suggesting that
progenitor cells had differentiated into postmitotic neurons upon inactivation
of Notch signaling as previously reported
(Austin et al., 1995
;
Henrique et al., 1997
).
Notably, Wnt2b largely suppressed the acute effect of DAPT to promote neuronal
differentiation; more than half of the cells still co-expressed Pax6 and Chx10
in the Wnt2b-expressing explants (Fig.
3H), and they incorporated BrdU as well
(Fig. 3L,U). The DAPT treatment
increased the number of visinin- or Hu-expressing cells in the
Wnt2b-expressing explants (Fig.
3C,D), suggesting that a certain population of cells was sensitive
to the inhibition of Notch signaling. To further confirm that Wnt2b can
suppress the premature neurogenesis caused by an inactivation of Notch
signaling, we co-expressed a dominant-negative form of Delta
(Henrique et al., 1997
) and
Wnt2b in the explants. For this experiment, we used two types of provirus
vector RCASBP (A) and (B) expressing different envelopes, which enables a
double infection of the same cell with two retroviruses
(Givol et al., 1994
). In the
retina co-electroporated with Delta1- and Wnt2b-expressing proviruses, both of
the genes were uniformly expressed, as was confirmed by in situ hybridization
on adjacent sections (data not shown). As previously reported
(Henrique et al., 1997
), the
dominant-negative form of Delta1 drastically increased the cells expressing
the neuronal marker (Fig. 3N).
This effect was completely neutralized by co-electroporation of Wnt2b
(Fig. 3P). In addition, Wnt2b
suppressed the effect of Delta1 to induce Müller glia differentiation
when co-expressed in the same explants
(Fig. 3Q-T). Those results
supported our hypothesis that Wnt2b inhibited progenitor cell differentiation
through a pathway independent of Delta/Notch signaling.
Wnt2b downregulates the expression of multiple proneural genes
To explore how Wnt2b could inhibit progenitor cell differentiation in the
absence of Notch activity, we studied the expression pattern of proneural
genes that are essential for the differentiation of retinal neurons (reviewed
by Vetter and Brown, 2001;
Perron and Harris, 2000a
;
Kageyama et al., 1997
). Before
that, we first examined the expression pattern of Notch1 mRNA itself
in retinas expressing Wnt2b using in situ hybridization. As previously
reported (Austin et al., 1995
),
Notch1 mRNA was broadly expressed in the undifferentiated cells of
the E5 neural retina (Fig. 4A).
The expression, however, was significantly reduced in the retina expressing
Wnt2b (Fig. 4B), which was
consistent with the aforementioned observation that Wnt2b-expressing retinal
explants became less sensitive to the treatment that blocks Notch signaling
(Fig. 3). We then examined the
expression pattern of atonal family proneural genes Cath5 and
NeuroM in adjacent sections
(Roztocil et al., 1997
;
Liu et al., 2001
). We observed
that the expression of both proneural genes was reduced in the
Wnt2b-expressing retina compared with that in the control retina
(Fig. 4C-F). Futhermore,
Delta1 expression was detected at the same level in both control and
Wnt2b-expressing retina (Fig.
4G,H). To further study the effect of Wnt2b on multiple proneural
gene expressions, we carried out RT-PCR using mRNAs derived from control or
Wnt2b-expressing E5 retina. We found that all the proneural genes we studied,
including Cath5, NeuroM, Cash1, Ngn1 and Ngn2, were
downregulated in the retina expressing Wnt2b
(Fig. 4I,J), while the
expression of Delta1 did not change
(Fig. 4I,J). In contrast, the
expression of Lef1 was upregulated in response to the Wnt2b
overexpression (Fig. 4I), as
previously reported (Kubo et al.,
2003
).
Exogenous expression of Cath5 suppresses the inhibitory effect of Wnt2b on retinal ganglion cell differentiation
The observations so far described suggested that Wnt2b inhibited progenitor
cell differentiation by negatively regulating the expression of proneural
genes. To confirm this, we asked if forced expression of a proneural gene
using an exogenous promoter could rescue the inhibitory effect of Wnt2b on
cellular differentiation. We focused on differentiation of retinal ganglion
cells, since molecular mechanisms leading to differentiation of this cell type
had been well characterized by a number of previous studies (reviewed by
Mu and Klein, 2004). The
atonal family bHLH transcription factor Ath5 (Cath5 in chicken, Math5 in
mouse, and Xash5 in frog) is the primary factor that specifies the retinal
ganglion cell lineage, and it activates transcription of other genes essential
for terminal differentiation of the cells (reviewed by
Mu and Klein, 2004
). We sought
to overexpress Cath5 and/or Wnt2b at early stages such as E2 and analyze at
E3.5, before the emergence of a huge number of endogenous retinal ganglion
cells that might obscure the exogenous ganglion cells induced by Cath5
overexpression. For this purpose, we overexpressed exogenous genes under the
control of the strong ubiquitous promoter CAG
(Niwa et al., 1991
), the
expression of which starts earlier than that of the retrovirus promoter we
used in the prior experiments mentioned above. As previously reported
(Liu et al., 2001
;
Matter-Sadzinski et al., 2001
;
Kanekar et al., 1997
),
overexpression of Cath5 increased the number of retinal ganglion cells
compared with their number in the control embryo
(Fig. 5B,F,I). On the contrary,
Wnt2b overexpression completely inhibited differentiation of retinal ganglion
cells (Fig. 5C,G,I). When Wnt2b
and Cath5 were co-introduced, differentiation of the retinal ganglion cells
was rescued in the cells expressing co-electroporated GFP
(Fig. 5D,H,I). These results
indicated that downregulation of proneural genes was necessary for Wnt2b to
inhibit progenitor cell differentiation.
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Discussion |
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Wnt activates diverse signaling pathways depending on the cell type
(reviewed by Wodarz and Nusse,
1998). Since Wnt2b stabilizes a cytosolic fraction of
ß-catenin in embryonic retinal cells
(Kubo et al., 2003
), it is
likely that Wnt2b stimulates the canonical Wnt pathway in the retina, which
activates target gene transcriptions through a downstream effector
ß-catenin/Lef (Tcf) complex (reviewed by
Wodarz and Nusse, 1998
).
Nevertheless, we observed downregulation of expression of multiple proneural
gene as well as Notch in the Wnt2b-overexpressed retina. Recently, the
ß-catenin/Lef (Tcf) complex has been shown to bind an upstream regulatory
region of Ngn1, a proneural bHLH gene essential for neuronal differentiation,
and to activate its transcription
(Hirabayashi et al., 2004
).
Interestingly, the activation of the canonical Wnt pathway promotes the
differentiation of cultured neural precursor cells derived from the E13.5
mouse cortex or E11.5 telencephalon, but inhibits their differentiation when
the precursor cells are prepared from E10.5 embryo
(Hirabayashi et al., 2004
;
Muroyama et al., 2004
).
Therefore, the ß-catenin/Lef (Tcf) complex may function both as a
repressor and an activator of the same target genes such as Ngn1 to exert
opposite effects, depending on the cellular context. Alternatively, Wnt2b may
activate the expression of certain transcriptional repressor(s), which in turn
downregulates the expression of target proneural genes in the retina
(Fig. 7A). Since an activation
of Notch signaling leads to glial cell differentiation
(Furukawa et al., 2000
;
Satow et al., 2001
;
Scheer et al., 2001
), it is
unlikely that Wnt2b stimulates Notch-downstream effector genes such as Hes to
inhibit proneural gene functions. Several studies have indicated the
importance of the exit from the cell cycle to the proper control of
differentiation (reviewed by Ohnuma and
Harris, 2003
), suggesting that Wnt2b might inhibit cellular
differentiation through its mitogenic activity, which promotes continuous cell
cycle progression. However, we have demonstrated that Wnt2b can inhibit
ganglion cell differentiation even when the cell cycle progression is
perturbed by Ink4D overexpression. We thus favor a model in which Wnt2b exerts
its negative effect on differentiation independently of its mitogenic
activity, presumably by negatively regulating proneural genes.
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ACKNOWLEDGMENTS |
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Footnotes |
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REFERENCES |
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Ahmad, I., Dooley, C. M. and Polk, D. L. (1997). Delta-1 is a regulator of neurogenesis in the vertebrate retina. Dev. Biol. 185,92 -103.[CrossRef][Medline]
Ahmad, I., Tang, L. and Pham, H. (2000). Identification of neural progenitors in the adult mammalian eye. Biochem. Biophys. Res. Commun. 270,517 -521.[CrossRef][Medline]
Austin, C. P., Feldman, D. E., Ida, J. A., Jr and Cepko, C.
L. (1995). Vertebrate retinal ganglion cells are selected
from competent progenitors by the action of Notch.
Development 121,3637
-3650.
Bao, Z. Z. and Cepko, C. L. (1997). The
expression and function of Notch pathway genes in the developing rat eye.
J. Neurosci. 17,1425
-1434.
Bard, J. B. and Ross, A. S. (1982). The morphogenesis of the ciliary body of the avian eye. I. Lateral cell detachment facilitates epithelial folding. Dev. Biol. 92, 73-86.[CrossRef][Medline]
Belecky-Adams, T., Tomarev, S., Li, H. S., Ploder, L., McInnes, R. R., Sundin, O. and Adler, R. (1997). Pax-6, Prox 1, and Chx10 homeobox gene expression correlates with phenotypic fate of retinal precursor cells. Invest. Ophthalmol. Vis. Sci. 38,1293 -1303.[Abstract]
Brown, N. L., Kanekar, S., Vetter, M. L., Tucker, P. K., Gemza,
D. L. and Glaser, T. (1998). Math5 encodes a murine basic
helix-loop-helix transcription factor expressed during early stages of retinal
neurogenesis. Development
125,4821
-4833.
Brown, N. L., Patel, S., Brzezinski, J. and Glaser, T.
(2001). Math5 is required for retinal ganglion cell and optic
nerve formation. Development
128,2497
-2508.
Chenn, A. and Walsh, C. A. (2002). Regulation
of cerebral cortical size by control of cell cycle exit in neural precursors.
Science 297,365
-369.
Cook, B., Portera-Cailliau, C. and Adler, R. (1998). Developmental neuronal death is not a universal phenomenon among cell types in the chick embryo retina. J. Comp. Neurol. 396,12 -19.[CrossRef][Medline]
Cunningham, J. J., Levine, E. M., Zindy, F., Goloubeva, O., Roussel, M. F. and Smeyne, R. J. (2002). The cyclin-dependent kinase inhibitors p19(Ink4d) and p27(Kip1) are coexpressed in select retinal cells and act cooperatively to control cell cycle exit. Mol. Cell. Neurosci. 19,359 -374.[CrossRef][Medline]
Dorsky, R. I., Rapaport, D. H. and Harris, W. A. (1995). Xotch inhibits cell differentiation in the Xenopus retina. Neuron 14,487 -496.[CrossRef][Medline]
Dorsky, R. I., Chang, W. S., Rapaport, D. H. and Harris, W. A. (1997). Regulation of neuronal diversity in the Xenopus retina by Delta signalling. Nature 385, 67-70.[CrossRef][Medline]
Dovey, H. F., John, V., Anderson, J. P., Chen, L. Z., de Saint Andrieu, P., Fang, L. Y., Freedman, S. B., Folmer, B., Goldbach, E., Holsztynska, E. J. et al. (2001). Functional gamma-secretase inhibitors reduce beta-amyloid peptide levels in brain. J. Neurochem. 76,173 -181.[CrossRef][Medline]
Dyer, M. A. and Cepko, C. L. (2001). Regulating proliferation during retinal development. Nat. Rev. Neurosci. 2,333 -342.[CrossRef][Medline]
Fischer, A. J. and Reh, T. A. (2000). Identification of a proliferating marginal zone of retinal progenitors in postnatal chickens. Dev. Biol. 220,197 -210.[CrossRef][Medline]
Fischer, A. J., Dierks, B. D. and Reh, T. A.
(2002). Exogenous growth factors induce the production of
ganglion cells at the retinal margin. Development
129,2283
-2291.
Fuhrmann, S., Levine, E. M. and Reh, T. A.
(2000). Extraocular mesenchyme patterns the optic vesicle during
early eye development in the embryonic chick.
Development 127,4599
-4609.
Furukawa, T., Mukherjee, S., Bao, Z. Z., Morrow, E. M. and Cepko, C. L. (2000). rax, Hes1, and notch1 promote the formation of Muller glia by postnatal retinal progenitor cells. Neuron 26,383 -394.[CrossRef][Medline]
Geling, A., Steiner, H., Willem, M., Bally-Cuif, L. and Haass,
C. (2002). A gamma-secretase inhibitor blocks Notch signaling
in vivo and causes a severe neurogenic phenotype in zebrafish. EMBO
Rep. 3,688
-694.
Givol, I., Tsarfaty, I., Resau, J., Rulong, S., da Silva, P. P., Nasioulas, G., DuHadaway, J., Hughes, S. H. and Ewert, D. L. (1994). Bcl-2 expressed using a retroviral vector is localized primarily in the nuclear membrane and the endoplasmic reticulum of chicken embryo fibroblasts. Cell Growth Differ. 5, 419-429.[Abstract]
Haruta, M., Kosaka, M., Kanegae, Y., Saito, I., Inoue, T., Kageyama, R., Nishida, A., Honda, Y. and Takahashi, M. (2001). Induction of photoreceptor-specific phenotypes in adult mammalian iris tissue. Nat. Neurosci. 4,1163 -1164.[CrossRef][Medline]
Hatakenaka, S., Kiyama, H., Tohyama, M. and Miki, N. (1985). Immunohistochemical localization of chick retinal 24 kdalton protein (visinin) in various vertebrate retinae. Brain Res. 331,209 -215.[CrossRef][Medline]
Henrique, D., Hirsinger, E., Adam, J., le Roux, I., Pourquie, O., Ish-Horowicz, D. and Lewis, J. (1997). Maintenance of neuroepithelial progenitor cells by Delta-Notch signalling in the embryonic chick retina. Curr. Biol. 7, 661-670.[CrossRef][Medline]
Hirabayashi, Y., Itoh, Y., Tabata, H., Nakajima, K., Akiyama,
T., Masuyama, N. and Gotoh, Y. (2004). The Wnt/{beta}-catenin
pathway directs neuronal differentiation of cortical neural precursor cells.
Development 131,2791
-2801.
Hollyfield, J. G. (1968). Differential addition of cells to the retina in Rana pipiens tadpoles. Dev. Biol. 18,163 -179.[CrossRef][Medline]
Hsieh, J. C., Rattner, A., Smallwood, P. M. and Nathans, J.
(1999). Biochemical characterization of Wnt-frizzled interactions
using a soluble, biologically active vertebrate Wnt protein. Proc.
Natl. Acad. Sci. USA 96,3546
-3551.
Hutcheson, D. A. and Vetter, M. L. (2001). The bHLH factors Xath5 and XNeuroD can upregulate the expression of XBrn3d, a POU-homeodomain transcription factor. Dev. Biol. 232,327 -338.[CrossRef][Medline]
Inoue, T., Hojo, M., Bessho, Y., Tano, Y., Lee, J. E. and Kageyama, R. (2002). Math3 and NeuroD regulate amacrine cell fate specification in the retina. Development 129,831 -842.[Medline]
Jacobson, M. (1968). Cessation of DNA synthesis in retinal ganglion cells correlated with the time of specification of their central conections. Dev. Biol. 17,219 -232.[Medline]
Jasoni, C., Hendrickson, A. and Roelink, H. (1999). Analysis of chicken Wnt-13 expression demonstrates coincidence with cell division in the developing eye and is consistent with a role in induction. Dev. Dyn. 215,215 -224.[CrossRef][Medline]
Johns, P. R. (1977). Growth of the adult goldfish eye. III. Source of the new retinal cells. J. Comp. Neurol. 176,343 -357.[CrossRef][Medline]
Kageyama, R., Ishibashi, M., Takebayashi, K. and Tomita, K. (1997). bHLH transcription factors and mammalian neuronal differentiation. Int. J. Biochem. Cell Biol. 29,1389 -1399.[CrossRef][Medline]
Kageyama, R., Ohtsuka, T. and Tomita, K. (2000). The bHLH gene Hes1 regulates differentiation of multiple cell types. Mol. Cells 10, 1-7.[CrossRef][Medline]
Kahn, A. J. (1974). An autoradiographic analysis of the time of appearance of neurons in the developing chick neural retina. Dev. Biol. 38,30 -40.[CrossRef][Medline]
Kanekar, S., Perron, M., Dorsky, R., Harris, W. A., Jan, L. Y., Jan, Y. N. and Vetter, M. L. (1997). Xath5 participates in a network of bHLH genes in the developing Xenopus retina. Neuron 19,981 -994.[CrossRef][Medline]
Kay, J. N., Finger-Baier, K. C., Roeser, T., Staub, W. and Baier, H. (2001). Retinal ganglion cell genesis requires lakritz, a Zebrafish atonal Homolog. Neuron 30,725 -736.[CrossRef][Medline]
Kelley, M. W., Turner, J. K. and Reh, T. A. (1995). Regulation of proliferation and photoreceptor differentiation in fetal human retinal cell cultures. Invest. Ophthalmol. Vis. Sci. 36,1280 -1289.[Abstract]
Kubo, F., Takeichi, M. and Nakagawa, S. (2003).
Wnt2b controls retinal cell differentiation at the ciliary marginal zone.
Development 130,587
-598.
Liu, H., Mohamed, O., Dufort, D. and Wallace, V. A. (2003). Characterization of Wnt signaling components and activation of the Wnt canonical pathway in the murine retina. Dev. Dyn. 227,323 -334.[CrossRef][Medline]
Liu, W., Mo, Z. and Xiang, M. (2001). The Ath5
proneural genes function upstream of Brn3 POU domain transcription factor
genes to promote retinal ganglion cell development. Proc. Natl.
Acad. Sci. USA 98,1649
-1654.
Livesey, F. J. and Cepko, C. L. (2001). Vertebrate neural cell-fate determination: lessons from the retina. Nat. Rev. Neurosci. 2,109 -118.[CrossRef][Medline]
Marquardt, T., Ashery-Padan, R., Andrejewski, N., Scardigli, R., Guillemot, F. and Gruss, P. (2001). Pax6 is required for the multipotent state of retinal progenitor cells. Cell 105, 43-55.[CrossRef][Medline]
Matter-Sadzinski, L., Matter, J. M., Ong, M. T., Hernandez, J.
and Ballivet, M. (2001). Specification of neurotransmitter
receptor identity in developing retina: the chick ATH5 promoter integrates the
positive and negative effects of several bHLH proteins.
Development 128,217
-231.
Megason, S. G. and McMahon, A. P. (2002). A
mitogen gradient of dorsal midline Wnts organizes growth in the CNS.
Development 129,2087
-2098.
Morrow, E. M., Furukawa, T., Lee, J. E. and Cepko, C. L.
(1999). NeuroD regulates multiple functions in the developing
neural retina in rodent. Development
126, 23-36.
Moshiri, A. and Reh, T. A. (2004). Persistent
progenitors at the retinal margin of ptc+/ mice. J.
Neurosci. 24,229
-237.
Mu, X. and Klein, W. H. (2004). A gene regulatory hierarchy for retinal ganglion cell specification and differentiation. Semin. Cell Dev. Biol. 15,115 -123.[CrossRef][Medline]
Muroyama, Y., Kondoh, H. and Takada, S. (2004). Wnt proteins promote neuronal differentiation in neural stem cell culture. Biochem. Biophys. Res. Commun. 313,915 -921.[CrossRef][Medline]
Nakagawa, S., Takada, S., Takada, R. and Takeichi, M. (2003). Identification of the laminar-inducing factor: Wnt-signal from the anterior rim induces correct laminar formation of the neural retina in vitro. Dev. Biol. 260,414 -425.[CrossRef][Medline]
Niwa, H., Yamamura, K. and Miyazaki, J. (1991). Efficient selection for high-expression transfectants with a novel eukaryotic vector. Gene 108,193 -199.[CrossRef][Medline]
Norenberg, M. D., Dutt, K. and Reif-Lehrer, L. (1980). Glutamine synthetase localization in cortisol-induced chick embryo retinas. J. Cell Biol. 84,803 -807.[Abstract]
Ohnuma, S. and Harris, W. A. (2003). Neurogenesis and the cell cycle. Neuron 40,199 -208.[CrossRef][Medline]
Ohnuma, S., Hopper, S., Wang, K. C., Philpott, A. and Harris, W. A. (2002). Co-ordinating retinal histogenesis: early cell cycle exit enhances early cell fate determination in the Xenopus retina. Development 129,2435 -2446.[Medline]
Perron, M. and Harris, W. A. (2000a). Determination of vertebrate retinal progenitor cell fate by the Notch pathway and basic helix-loop-helix transcription factors. Cell Mol. Life Sci. 57,215 -223.[Medline]
Perron, M. and Harris, W. A. (2000b). Retinal stem cells in vertebrates. BioEssays 22,685 -688.[CrossRef][Medline]
Perron, M., Kanekar, S., Vetter, M. L. and Harris, W. A. (1998). The genetic sequence of retinal development in the ciliary margin of the Xenopus eye. Dev. Biol. 199,185 -200.[CrossRef][Medline]
Perron, M., Opdecamp, K., Butler, K., Harris, W. A. and
Bellefroid, E. J. (1999). X-ngnr-1 and Xath3 promote ectopic
expression of sensory neuron markers in the neurula ectoderm and have distinct
inducing properties in the retina. Proc. Natl. Acad. Sci.
USA 96,14996
-15001.
Reh, T. A. and Levine, E. M. (1998). Multipotential stem cells and progenitors in the vertebrate retina. J. Neurobiol. 36,206 -220.[CrossRef][Medline]
Roztocil, T., Matter-Sadzinski, L., Alliod, C., Ballivet, M. and
Matter, J. M. (1997). NeuroM, a neural helix-loop-helix
transcription factor, defines a new transition stage in neurogenesis.
Development 124,3263
-3272.
Satow, T., Bae, S. K., Inoue, T., Inoue, C., Miyoshi, G.,
Tomita, K., Bessho, Y., Hashimoto, N. and Kageyama, R.
(2001). The basic helix-loop-helix gene hesr2 promotes
gliogenesis in mouse retina. J. Neurosci.
21,1265
-1273.
Scheer, N., Groth, A., Hans, S. and Campos-Ortega, J. A.
(2001). An instructive function for Notch in promoting
gliogenesis in the zebrafish retina. Development
128,1099
-1107.
Schmidt, M., Tanaka, M. and Munsterberg, A.
(2000). Expression of (beta)-catenin in the developing chick
myotome is regulated by myogenic signals. Development
127,4105
-4113.
Sidman, R. (1961). Histogenesis of Mouse Retina Studied with Thymidine-H3. New York: Academic Press.
Tetsu, O. and McCormick, F. (1999). Beta-catenin regulates expression of cyclin D1 in colon carcinoma cells. Nature 398,422 -426.[CrossRef][Medline]
Thut, C. J., Rountree, R. B., Hwa, M. and Kingsley, D. M. (2001). A large-scale in situ screen provides molecular evidence for the induction of eye anterior segment structures by the developing lens. Dev. Biol. 231,63 -76.[CrossRef][Medline]
Tomita, K., Nakanishi, S., Guillemot, F. and Kageyama, R.
(1996). Mash1 promotes neuronal differentiation in the retina.
Genes Cells 1,765
-774.
Tomita, K., Moriyoshi, K., Nakanishi, S., Guillemot, F. and
Kageyama, R. (2000). Mammalian achaete-scute and atonal
homologs regulate neuronal versus glial fate determination in the central
nervous system. EMBO J.
19,5460
-5472.
Tropepe, V., Coles, B. L., Chiasson, B. J., Horsford, D. J.,
Elia, A. J., McInnes, R. R. and van der Kooy, D. (2000).
Retinal stem cells in the adult mammalian eye. Science
287,2032
-2036.
Vetter, M. L. and Brown, N. L. (2001). The role of basic helix-loop-helix genes in vertebrate retinogenesis. Semin. Cell Dev. Biol. 12,491 -498.[CrossRef][Medline]
Wang, S. W., Kim, B. S., Ding, K., Wang, H., Sun, D., Johnson,
R. L., Klein, W. H. and Gan, L. (2001). Requirement for math5
in the development of retinal ganglion cells. Genes
Dev. 15,24
-29.
Wodarz, A. and Nusse, R. (1998). Mechanisms of Wnt signaling in development. Annu. Rev. Cell Dev. Biol. 14,59 -88.[CrossRef][Medline]
Yan, R. T. and Wang, S. Z. (1998). neuroD induces photoreceptor cell overproduction in vivo and de novo generation in vitro. J. Neurobiol. 36,485 -496.[CrossRef][Medline]
Zakin, L. D., Mazan, S., Maury, M., Martin, N., Guenet, J. L. and Brulet, P. (1998). Structure and expression of Wnt13, a novel mouse Wnt2 related gene. Mech. Dev. 73,107 -116.[CrossRef][Medline]