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First published online December 20, 2005
doi: 10.1242/10.1242/dev.02208
1 Department of Cell and Molecular Biology, Medical Nobel Institute, Karolinska
Institute, SE-171 77 Stockholm, Sweden.
2 Department of Biotechnology, The Royal Institute of Technology, SE-106 91
Stockholm, Sweden.
3 Department of Oncology-Pathology, Karolinska Institute, SE-171 76, Stockholm,
Sweden.
* Author for correspondence (e-mail: jonas.frisen{at}cmb.ki.se)
Accepted 10 November 2005
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SUMMARY |
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 TOP SUMMARY INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Key words: p53, Self-renewal, Stem cell, p21 (Cdkn1a), Adult, Cancer, Mouse, Trp53, Microarray data
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INTRODUCTION |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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).
Several proto-oncogenes and tumor suppressors, such as Bmi1
(Molofsky et al., 2003), Gfi1
(Hock et al., 2004) and Pten
(Groszer et al., 2001), control
the self-renewal of normal untransformed tissue stem cells. This indicates
that key components of the molecular regulation of tissue and cancer stem cell
features may be shared, and that tumor formation, in at least some aspects,
can be viewed as excessive stem cell expansion
(Pardal et al., 2003). There
is indeed evidence that some tumors may originate from tissue stem cells,
whereas other may arise by the dedifferentiation of progenitor cells to a more
stem cell like state (Daley,
2004). The self-renewal of tissue and cancer stem cells can be
regulated by modulation of several distinct processes, including cell
proliferation, cell death and differentiation
(Molofsky et al., 2004).
Understanding the molecular pathways controlling stem cell self-renewal may
shed light on both tissue homeostasis and cancer development and
progression.
p53 (Trp53 - Mouse Genome Informatics; TP53 - Human Gene Nomenclature
Database) is the prototypical tumor suppressor gene and this pathway is
inactivated in most human cancers
(Vogelstein et al., 2000). In
spite of its key role in tumor development, a potential function for p53 in
tissue stem cells has not been addressed. p53 mutations are common in brain
tumors, and are implicated in both tumor initiation and growth
(Sidransky et al., 1992). The
identification of cancer stem cells in brain tumors suggests that molecular
mechanisms controlling neural stem cell proliferation and brain tumor
initiation and growth may be shared (Singh
et al., 2004b). We have addressed the function of p53 in neural
stem cells in the adult brain, and report that it acts as a negative regulator
of neural stem cell self-renewal.
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MATERIALS AND METHODS |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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) on a C57BL/6 background (8-12 weeks old) were used with
their corresponding littermate wild-type controls. At least four animals of
each genotype were used for all experiments.
Immunohistochemistry
All incubations were performed in a humidified chamber for 16 hours at
4°C overnight. Citrate antigen retrieval was used for p53 protein
detection by boiling sections for 15 minutes in a 10 mM citrate solution at pH
6. For signal amplification, we used a combination of ABC staining (Vector
Laboratories) and TSA (Perkin Elmer). The following primary antibodies were
used: anti-phospo-histone H3 (1:2000, Upstate), anti-p53 (1:500, Novocastra),
anti-Gfap (1:2000, DakoCytomation), anti-ßIII-tubulin (Tuj1, 1:1000,
Biosite), anti-O4 (1:100, Chemicon), anti-BrdU (1:1000, Chemicon), anti-Dcx
(1:1000, Chemicon) and anti-musashi 1 (1:500, gift from H. Okano). Images were
acquired on a Zeiss LSM510 Meta confocal microscope and a Zeiss Axioplan2 with
Openlab software.
Apoptosis detection
Apoptotic cells with fragmented DNA were detected in tissue sections using
the FragEL TUNEL kit (Calbiochem). In vitro apoptosis was detected by Annexin
V (BD) labeling on single cells dissociated from small neurospheres 4 hours
prior to staining or caspase activity using the Vybrant FAM assay (Molecular
Probes), and both analyzed with a FACScalibur (BD) flowcytometer.
Neural stem cell cultures
The lateral walls of the lateral ventricles were dissected out and cells
dissociated using papain (Worthington). Neurospheres were cultured as
described (Johansson et al.,
1999) in DMEM/F12 medium supplemented with B27 and EGF and bFGF
(both 10 ng/ml). Cells were passaged every 4th day by dissociating
neurospheres into single cells with papain. All experiments were performed
between passage 2 and 5. Neural stem cell differentiation was induced on
Biocoat polylysine/laminin culture slides with the addition of 1% calf serum
for 7 days before fixing in 4% formaldehyde in PBS and processing for
immunohistochemistry. Photos for size comparisons were taken 6 days after
clonal expansion of a single cell.
Self-renewal assay
Neurospheres from p53-/- and p53+/+ mice
were dissociated into single cells with papain, cell profiles were compared by
FACS, cell numbers counted and plated at clonal density. The number of
secondary neurospheres generated from 2000 single cells in triplicate from
four mice of each genotype was quantified after 7 days. Cells from the clonal
expansion were used for assaying multipotency by in vitro differentiation and
for photos of neurosphere size. Neurosphere photos for size comparisons were
taken 6 days after clonal expansion of a single cell. Primary neurosphere
formation from adult lateral ventricle wall was assessed by sorting single
cells into 96-well plates using FACS (BD, FACSVantage SE Diva) and counting
neurospheres 7 days later.
Proliferation assays
BrdU labeling of proliferating cells in vivo was performed by three
intraperitoneal injections of 50 mg/kg BrdU with 2 hours intervals and
followed by sacrifice 2 hours after the last injection. Anesthetized mice were
perfused transcardially with 4% formaldehyde in PBS and sectioned (25 µm)
on a vibratome (Leica).
Positive nuclei for BrdU or phosphorylated histone H3 were counted on serial coronal sections of the anterior horn of the lateral ventricle within a defined space of the subventricular zone. Sections were taken from coded animals and the genotype of the mice was revealed after statistical analysis.
For analysis in vitro, BrdU was added to cultures at a final concentration of 10 µM 30 minutes prior to fixation. BrdU detection was carried out with BrdU Flow (BD) on dissociated single cells and analyzed in a FACScalibur flowcytometer (BD).
Microarray analysis
Total RNA was prepared as two biological replicates on different occasions
from neurospheres using the RNeasy purification system (Qiagen). Amplification
of RNA was carried out using the RiboAmp total RNA amplification kit
(Arcturus). Each sample was hybridized against the reference sample using two
replicated hybridizations with the same dye assignments (reference sample
always in Cy3). Every cDNA probe was printed twice on an array, there were
therefore eight measurements in total for each genotype (two within-array
replicates, two replicated hybridizations and two replicated neurosphere
cultures). The reference sample consisted of Universal Reference RNA
(Stratagene) amplified using the same approach as the neurosphere samples.
Labeled material was hybridized to 16k mouse cDNA arrays using a
reference-design approach (Sievertzon et
al., 2005). All data analysis steps were carried out in the R
environment for statistical computing and programming using several analysis
packages as detailed in the supplementary material. Data corresponding to
unreliable features and non-expressed genes were removed, and the remaining
data normalized using an intensity-dependent print-tip lowess approach.
Differentially expressed genes were identified using an empirical Bayes
moderated t-test; a gene with a false-discovery rate adjusted
P<0.05 and an M>±0.4 (corresponds to a fold-change value
of greater than 1.4) was considered differentially expressed. Additional
details regarding the sample preparation, array hybridization and subsequent
data analysis are available on request. Additional information on the cDNA
microarray used can be obtained from ArrayExpress microarray data repository
(Accession number E-MEXP-483). Further details on the array procedure can be
provided on request.
Real-time PCR
RNA was isolated with Cells-to-Signal Lysis Buffer (Ambion) and cDNA was
synthesized with Superscript and random hexamers (Invitrogen). Linear phase of
logarithmic amplification was used for quantification and cycle number was
compared between triplicate samples. SYBR green was used on an ABI machine
according to the manufacturer's instructions.
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RESULTS |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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), but it has not been established which cell types express
p53. Moreover, p53 is subject to post-translational regulation, so mRNA levels
do not always accurately reflect protein levels
(Bode and Dong, 2004). Nuclear p53-immunoreactivity was evident in the majority of cells in the lateral ventricle wall neural stem cell niche (Fig. 1). Most cells in surrounding brain regions displayed much weaker or undetectable labeling. All labeling was abolished in sections from p53-null mice, corroborating the specificity of the antibody (data not shown). Multiciliated ependymal cells lining the ventricle appeared uniformly p53-immunoreactive (Fig. 1). The majority of cells in the subventricular zone, including most Gfap+ astrocytes and musashi 1+ progenitor cells displayed p53-immunoreactivity (Fig. 1A-E,G,H). p53 immunoreactivity was weaker or absent in the majority of differentiating cells, including Doublecortin+ neuroblasts (Dcx in Fig. 1F), in the migration path to the olfactory bulb (the rostral migratory stream). Thus, p53 protein is present at substantially higher levels in the neural stem cell lineage than in other cells in the adult brain, and is mainly detected in candidate stem cells and progenitor cells.
Increased cell proliferation in the neural stem cell niche in the absence of p53
To assess a potential role for p53 in the adult neural stem cell lineage,
we first analyzed the cell proliferation and apoptosis rate in the lateral
ventricle wall in mice lacking p53
(Donehower et al., 1992). We
found that p53-/- mice have a significantly higher number
of proliferating cells, as assessed by the incorporation of the nucleotide
analogue BrdU, in the lateral ventricle wall compared to wild-type littermates
(Fig. 2A-C). This was
paralleled by a similar increase in the number of cells immunoreactive to
phosphorylated Ser10 on histone 3, a mitosis-specific marker
(Wei et al., 1999), in the
mutant mice (Fig. 2D-F). The
distribution of proliferating cells in the subventricular zone along the
lateral ventricle wall and in the rostral migratory stream was
indistinguishable between wild-type and p53-null mice.
|
), and we
could not detect any significant difference between wild-type and
p53-null mice (Fig.
2G), making it unlikely that the increase in proliferation
is due to a decrease in cell death.
Negative regulation of neural stem cell self-renewal
Multipotent self-renewing neural stem cells can be propagated in vitro as
clonal aggregates denoted neurospheres
(Reynolds and Weiss, 1992).
Although these cells display hallmark stem cell properties in vitro, it is
unclear whether the neurosphere-forming cells act as stem and/or progenitor
cells in vivo. We asked whether the increased proliferation in the lateral
ventricle wall of p53-/- mice was accompanied by an
alteration in the number of cells capable of forming neurospheres. Indeed, a
significantly larger proportion of lateral ventricle wall cells from
p53-/- mice formed neurospheres compared with wild-type
cells (Fig. 2H).
Cells from p53-/- animals generated significantly larger neurospheres compared with wild-type cells (Fig. 3A-C). This was not due to a difference in cell size (Fig. 3L) but to a larger number of cells in the p53 null neurospheres.
Neurospheres are initiated by multipotent neural stem cells, but as the clone expands, an increasing heterogeneity will ensue with many cells committing to specific fates. Therefore, it cannot be directly inferred that an increase in cell number in a neurosphere is due to increased stem cell self-renewal. To directly assay the number of stem cells in the neurospheres, we analyzed the number of cells that were capable of forming new secondary neurospheres. This revealed a higher number of neurosphere-initiating cells in p53-null neurospheres, establishing that there was an increased generation of cells with in vitro neural stem cell potential in the absence of p53. By analyzing the relative frequency of neurosphere cells that were capable of reinitiating a clone, we found that this was not only due to the larger number of cells in these neurospheres, but the proportion of cells that were able to reinitiate neurosphere formation was significantly higher in p53-/- compared with wild-type neurospheres (Fig. 3D). The increased number of neural stem cells generated in the absence of p53 establishes p53 as a negative regulator of neural stem cell self-renewal.
p53 regulates neural stem cell proliferation and apoptosis
Stem cell self-renewal can be regulated by the modulation of any process
that affects the number of progeny from a stem cell maintaining stem cell
properties, including for example cell proliferation, survival or
differentiation (Mikkers and
Frisén, 2005; Molofsky
et al., 2004). To address by which mechanism p53 regulates neural
stem cell self-renewal, we assessed these parameters in p53 null and wild-type
neurospheres. Owing to the rapidly emerging size difference between p53 null
and wild-type neurospheres, we performed all analyses on small secondary
neurospheres (2 days after passage when the neurospheres contain 
4-20
cells), to minimize differences secondary to variation in neurosphere size. We
first analyzed proliferation by BrdU incorporation in neurospheres. As in the
lateral ventricle wall in vivo, BrdU incorporation was significantly increased
in vitro in neurospheres in the absence of p53
(Fig. 3J,K).
We next assessed apoptotic cell death. In contrast to the in vivo
situation, where very few apoptotic cells are seen
(Biebl et al., 2000), there is
a substantial proportion of apoptotic cells in neurospheres, which can be
detected by flowcytometric analysis of caspase activity or AnnexinV
immunoreactivity. We found that significantly fewer cells labeled with these
markers in p53-/- compared with wild-type neurospheres
(Fig. 3E,F), indicating
decreased apoptosis in the absence of p53. The phenotype appears more
pronounced in vitro than in vivo, with a larger increase in proliferation as
well as reduced apoptosis in p53-/- neural stem cells in
vitro. This may be related to the often increased p19Arf levels in
cultured cells (Molofsky et al.,
2005). As p19Arf is a positive regulator of p53
expression (Zindy et al.,
2003), the difference between wild-type and p53-null
cells may be augmented in vitro.
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Taken together, these data indicate that the increased self-renewal of neural stem cells in the absence of p53 is mediated by the combination of an increase in cell proliferation and a decrease in apoptotic cell death.
p53 regulation of the neural stem cell transcriptome
To gain insight into how p53 impinges on the molecular program controlling
self-renewal, we analyzed the transcriptome of p53-/- and
wild-type neurospheres using 16k cDNA arrays enriched for genes expressed in
murine stem cell populations (Sievertzon
et al., 2005). Cultured cells are likely to differ from their in
vivo counterparts as p53 regulation may be affected, but allow for a rather
homogeneous population to be analyzed. We used the empirical Bayes moderated
t-test to identify differential expression. The contribution of
within-array duplicate features was downweighted and genes with a
false-discovery rate adjusted P<0.05 combined with an M-value
cut-off (|M|>0.4) were considered differentially expressed
(details can be provided on request). Using these criteria, 325 genes were
differentially expressed and 98 of these had P<0.001
(Fig. 4A,
Table 1; see Table S1 in the
supplementary material for a complete list). The expression of seven of these
genes was previously reported to be controlled by p53
(Harms et al., 2004),
validating the approach. However, the majority of dysregulated genes have not
been implicated as p53 targets. Functional classification of dysregulated
genes revealed significant enrichment of several categories related to cell
proliferation (gene ontology themes with an Ease-score<0.05,
Fig. 4B; see Table S3 in the
supplementary material).
|
), and the
decreased expression of these genes in concert with altered expression of
other modulators of proliferation and apoptosis (see Table S1 in the
supplementary material) is likely to contribute to the observed phenotype in
the neurospheres. Western blot analysis of p21 protein levels in the lateral
ventricle wall revealed that p53-/- mice had 77±37%
(average±s.d., n=6 of each genotype) of the level seen in
wild-type mice (Fig. 4D). Several pathways implicated in stem cell self-renewal regulate p53 expression
or activity (Fig. 4E),
suggesting that p53 may be an important modulator of self-renewal in multiple
lineages.
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DISCUSSION |
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TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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The analysis of the transcriptional effects of loss of p53 in neurospheres
unveiled downstream candidates potentially responsible for this phenotype.
Although redundancy between different pathways often can mask a role of an
individual gene in the analysis of mutant mice, we found several dysregulated
genes in p53-/- neurospheres. There is also a risk of a
developmental phenotype in a mutant giving secondary effects in adulthood.
With the exception of a subset of p53-null mice developing neural
tube defects and dying at birth (Sah et
al., 1995), the nervous system appears normal in adult p53-null
mice. The most prominent dysregulation was the reduction of p21 in
p53-/- neurospheres. p21 is a well-established actor in
p53-mediated cell cycle arrest and through its negative effects on CDKs, p21
inhibits both the G1-to-S and the G2-to-mitosis transitions
(Gartel and Radhakrishnan,
2005). p21 negatively regulates self-renewal of hematopoietic stem
cells (Cheng et al., 2000) and
the precipitous decrease in expression in p53-/-
neurospheres may contribute to the increased self-renewal. Indeed,
proliferation is increased in the lateral ventricle wall and in neurospheres
in the absence of p21 in young adult mice
(Kippin et al., 2005). Another
study found increased proliferation in p21-/- mice only
after ischemia and not in the intact brain
(Qiu et al., 2004). Moreover,
p27 has been established as a negative regulator of proliferation in the
lateral ventricle wall in the adult brain
(Doetsch et al., 2002). The
decreased expression of cip/kip family genes in conjunction with the
upregulation of a large number of genes positively regulating cell
proliferation and downregulation of several pro-apoptotic genes (see Table S1
in the supplementary material) in p53-/- neural stem cells
is likely to contribute to the observed phenotype.
Kippin et al. recently demonstrated an exhaustion of neural stem cells in
old p21-/- mice
(Kippin et al., 2005). This
suggests that increased proliferation may result in premature senescence and
that it may be important to suppress self-renewal for the long-term
maintenance of a stem cell population. The limited life span of
p53-null mice because of their predisposition to tumors precludes an
analysis of the role of p53 in the aging of stem cells.
A recent study demonstrated that p53 promotes the differentiation of
embryonic stem cells by suppressing Nanog expression
(Lin et al., 2005). This
suggests that p53-mediated suppression of stem cell self-renewal may be
general. However, the mechanisms by which p53 regulate self-renewal are
different, as Nanog expression is restricted to embryonic stem cells and
germ-line stem cells.
Several recent studies have described the existence of a subpopulation of
cells in brain tumors that has characteristics of stem cells
(Singh et al., 2004a;
Singh et al., 2004b). The
concept that a cancer stem cell is maintaining the growth of a tumor has
important implications for our understanding of tumorigenesis and design of
cancer therapy. The cancer stem cell hypothesis suggests that cells in a tumor
have different abilities regarding proliferation, self-renewal and
differentiation (Singh et al.,
2004a). The influence of p53 inactivation on these processes is an
important factor to understand as p53 is mutated in the majority of tumors
found throughout the human body
(Vogelstein et al., 2000). The
mutations that inactivate p53 function in cancer cells almost all appear to
localize to the DNA-binding domain of the p53 protein and produce a protein
that fails to transcribe p53-responsive genes
(Harris and Levine, 2005). We
have in this study chosen to analyze the p53 pathway in adult neural stem and
progenitor cells in vivo and in vitro in order to gain insight into the
molecular mechanisms regulating proliferation and self-renewal. Focusing on
the proliferative zones of the adult brain for the study of p53 is due to the
suggested connection between tumorigenesis and sites of immature cells.
Furthermore, neural progenitor cells retain the ability to proliferate and
self-renew throughout adulthood and may be susceptible to convert into a
malignant phenotype and may serve as an interesting model for the cancer stem
cell hypothesis. The increased proliferation rate in the neural stem cell
niche may elevate the risk of acquiring mutations, which could contribute with
other previously described mechanisms
(Vogelstein et al., 2000) to
the increased predisposition to cancer in the absence of p53.
There is accumulating evidence for the role of tumor suppressors and
oncogenes in the maintenance and regulation of stem cells. Our array analysis
of neural stem cells lacking p53 provides an overview of the pathways that are
important for the increased self-renewal phenotype observed in
p53-/- neurospheres. When compared with previous array
studies of neural stem cells (Fortunel et
al., 2003), we find that few of the genes with enriched expression
in neural stem cells are dysregulated in the absence of p53 (see Fig. S1 in
the supplementary material). Instead, many of the dysregulated genes control
cell cycle progression and apoptosis without affecting differentiation. Our
molecular description of a self-renewal phenotype using a cDNA array based
analysis of the transcriptome opens up pathways for the discovery of genes
that regulate this fundamental property of stem cells. The functional analysis
of other known oncogenes and tumor suppressors in stem cell maintenance,
together with the present data, suggests that several molecular pathways in
stem cell biology may converge on p53 for the control of stem cell
self-renewal (Fig. 4E).
Supplementary material
Supplementary material for this article is available at
http://dev.biologists.org/cgi/content/full/133/2/363/DC1
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ACKNOWLEDGMENTS |
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|
REFERENCES |
|---|
|
TOPSUMMARYINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
|---|
Biebl, M., Cooper, C. M., Winkler, J. and Kuhn, H. G. (2000). Analysis of neurogenesis and programmed cell death reveals a self-renewing capacity in the adult rat brain. Neurosci. Lett. 291,17 -20.[CrossRef][Medline]
Bode, A. M. and Dong, Z. (2004). Post-translational modification of p53 in tumorigenesis. Nat. Rev. Cancer 4,793 -805.[CrossRef][Medline]
Cheng, T., Rodrigues, N., Dombkowski, D., Stier, S. and Scadden, D. T. (2000). Stem cell repopulation efficiency but not pool size is governed by p27(kip1). Nat. Med. 6,1235 -1240.[CrossRef][Medline]
Daley, G. Q. (2004). Chronic myeloid leukemia: proving ground for cancer stem cells. Cell 119,314 -316.[CrossRef][Medline]
Doetsch, F., Verdugo, J. M., Caille, I., Alvarez-Buylla, A.,
Chao, M. V. and Casaccia-Bonnefil, P. (2002). Lack of the
cell-cycle inhibitor p27Kip1 results in selective increase of
transit-amplifying cells for adult neurogenesis. J.
Neurosci. 22,2255
-2264.
Donehower, L. A., Harvey, M., Slagle, B. L., McArthur, M. J., Montgomery, C. A., Jr, Butel, J. S. and Bradley, A. (1992). Mice deficient for p53 are developmentally normal but susceptible to spontaneous tumours. Nature 356,215 -221.[CrossRef][Medline]
Fortunel, N. O., Otu, H. H., Ng, H. H., Chen, J., Mu, X., Chevassut, T., Li, X., Joseph, M., Bailey, C., Hatzfeld, J. A. et al. (2003). Comment on "`Stemness': transcriptional profiling of embryonic and adult stem cells" and "a stem cell molecular signature" (and author reply). Science 302, 393.
Gartel, A. L. and Radhakrishnan, S. K. (2005).
Lost in transcription: p21 repression, mechanisms, and consequences.
Cancer Res. 65,3980
-3985.
Groszer, M., Erickson, R., Scripture-Adams, D. D., Lesche, R.,
Trumpp, A., Zack, J. A., Kornblum, H. I., Liu, X. and Wu, H.
(2001). Negative regulation of neural stem/progenitor cell
proliferation by the Pten tumor suppressor gene in vivo.
Science 294,2186
-2189.
Harms, K., Nozell, S. and Chen, X. (2004). The common and distinct target genes of the p53 family transcription factors. Cell Mol. Life Sci. 61,822 -842.[CrossRef][Medline]
Harris, S. L. and Levine, A. J. (2005). The p53 pathway: positive and negative feedback loops. Oncogene 24,2899 -2908.[CrossRef][Medline]
Hock, H., Hamblen, M. J., Rooke, H. M., Schindler, J. W., Saleque, S., Fujiwara, Y. and Orkin, S. H. (2004). Gfi-1 restricts proliferation and preserves functional integrity of haematopoietic stem cells. Nature 431,1002 -1007.[CrossRef][Medline]
Johansson, C. B., Momma, S., Clarke, D. L., Risling, M., Lendahl, U. and Frisén, J. (1999). Identification of a neural stem cell in the adult mammalian central nervous system. Cell 96,25 -34.[CrossRef][Medline]
Kippin, T. E., Martens, D. J. and van der Kooy, D.
(2005). p21 loss compromises the relative quiescence of forebrain
stem cell proliferation leading to exhaustion of their proliferation capacity.
Genes Dev. 19,756
-767.
Lin, T., Chao, C., Saito, S., Mazur, S. J., Murphy, M. E., Appella, E. and Xu, Y. (2005). p53 induces differentiation of mouse embryonic stem cells by suppressing Nanog expression. Nat. Cell Biol. 7,165 -171.[CrossRef][Medline]
Mikkers, H. and Frisén, J. (2005). Deconstructing stemness. EMBO J. 24,2715 -2719.[CrossRef][Medline]
Molofsky, A. V., Pardal, R., Iwashita, T., Park, I. K., Clarke, M. F. and Morrison, S. J. (2003). Bmi-1 dependence distinguishes neural stem cell self-renewal from progenitor proliferation. Nature 425,962 -967.[CrossRef][Medline]
Molofsky, A. V., Pardal, R. and Morrison, S. J. (2004). Diverse mechanisms regulate stem cell self-renewal. Curr. Opin. Cell Biol. 16,700 -707.[CrossRef][Medline]
Molofsky, A. V., He, S., Bydon, M., Morrison, S. J. and Pardal,
R. (2005). Bmi-1 promotes neural stem cell self-renewal and
neural development but not mouse growth and survival by repressing the
p16Ink4a and p19Arf senescence pathways. Genes Dev.
19,1432
-1437.
Pardal, R., Clarke, M. F. and Morrison, S. J. (2003). Applying the principles of stem-cell biology to cancer. Nat. Rev. Cancer 3,895 -902.[CrossRef][Medline]
Qiu, J., Takagi, Y., Harada, J., Rodrigues, N., Moskowitz, M.
A., Scadden, D. T. and Cheng, T. (2004). Regenerative
response in ischemic brain restricted by p21cip1/Waf1. J. Exp.
Med. 199,937
-945.
Reynolds, B. A. and Weiss, S. (1992).
Generation of neurons and astrocytes from isolated cells of the adult
mammalian nervous system. Science
255,1707
-1710.
Sah, V. P., Attardi, L. D., Mulligan, G. J., Williams, B. O., Bronson, R. T. and Jacks, T. (1995). A subset of p53-deficient embryos exhibit exencephaly. Nat. Genet. 10,175 -180.[CrossRef][Medline]
Sidransky, D., Mikkelsen, T., Schwechheimer, K., Rosenblum, M. L., Cavanee, W. and Vogelstein, B. (1992). Clonal expansion of p53 mutant cells is associated with brain tumour progression. Nature 355,846 -847.[CrossRef][Medline]
Sievertzon, M., Wirta, V., Mercer, A., Meletis, K., Erlandsson, R., Wikstrom, L., Frisen, J. and Lundeberg, J. (2005). Transcriptome analysis in primary neural stem cells using a tag cDNA amplification method. BMC Neurosci. 6, 28.[Medline]
Singh, S. K., Clarke, I. D., Hide, T. and Dirks, P. B. (2004a). Cancer stem cells in nervous system tumors. Oncogene 23,7267 -7273.[CrossRef][Medline]
Singh, S. K., Hawkins, C., Clarke, I. D., Squire, J. A., Bayani, J., Hide, T., Henkelman, R. M., Cusimano, M. D. and Dirks, P. B. (2004b). Identification of human brain tumour initiating cells. Nature 432,396 -401.[CrossRef][Medline]
van Lookeren Campagne, M. and Gill, R. (1998). Tumor-suppressor p53 is expressed in proliferating and newly formed neurons of the embryonic and postnatal rat brain: comparison with expression of the cell cycle regulators p21Waf1/Cip1, p27Kip1, p57Kip2, p16Ink4a, cyclin G1, and the proto-oncogene Bax. J. Comp. Neurol. 397,181 -198.[CrossRef][Medline]
Vogelstein, B., Lane, D. and Levine, A. J. (2000). Surfing the p53 network. Nature 408,307 -310.[CrossRef][Medline]
Wei, Y., Yu, L., Bowen, J., Gorovsky, M. A. and Allis, C. D. (1999). Phosphorylation of histone H3 is required for proper chromosome condensation and segregation. Cell 97, 99-109.[CrossRef][Medline]
Zindy, F., Quelle, D. E., Roussel, M. F. and Sherr, C. J. (2003). Expression of the p16INK4a tumor suppressor versus other INK4family members during mouse development and aging. Oncogene 15,203 -211.
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 C. Q. Doe Neural stem cells: balancing self-renewal with differentiation Development, May 1, 2008; 135(9): 1575 - 1587. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K. E. Galvin, H. Ye, D. J. Erstad, R. Feddersen, and C. Wetmore Gli1 Induces G2/M Arrest and Apoptosis in Hippocampal but Not Tumor-Derived Neural Stem Cells Stem Cells, April 1, 2008; 26(4): 1027 - 1036. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. N. Pechnick, S. Zonis, K. Wawrowsky, J. Pourmorady, and V. Chesnokova p21Cip1 restricts neuronal proliferation in the subgranular zone of the dentate gyrus of the hippocampus PNAS, January 29, 2008; 105(4): 1358 - 1363. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Merlino and C. Khanna Fishing for the origins of cancer Genes & Dev., June 1, 2007; 21(11): 1275 - 1279. [Full Text] [PDF]
|
|
|
|
|
|
L. Soroceanu, S. Kharbanda, R. Chen, R. H. Soriano, K. Aldape, A. Misra, J. Zha, W. F. Forrest, J. M. Nigro, Z. Modrusan, et al. Identification of IGF2 signaling through phosphoinositide-3-kinase regulatory subunit 3 as a growth-promoting axis in glioblastoma PNAS, February 27, 2007; 104(9): 3466 - 3471. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. I. Kornblum Introduction to Neural Stem Cells Stroke, February 1, 2007; 38(2): 810 - 816. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Papers of Note Sci. Aging Knowl. Environ., January 4, 2006; 2006(1): nw1 - nw1. [Full Text]
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