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DNMT1 maintains progenitor function in self-renewing somatic tissue.

Sen GL, Reuter JA, Webster DE, Zhu L, Khavari PA - Nature (2010)

Bottom Line: DNMT1 protein was found enriched in undifferentiated cells, where it was required to retain proliferative stamina and suppress differentiation.Genome-wide analysis showed that a significant portion of epidermal differentiation gene promoters were methylated in self-renewing conditions but were subsequently demethylated during differentiation.These data demonstrate that proteins involved in the dynamic regulation of DNA methylation patterns are required for progenitor maintenance and self-renewal in mammalian somatic tissue.

View Article: PubMed Central - PubMed

Affiliation: Programs in Epithelial Biology and Cancer Biology and the Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California 94305, USA.

ABSTRACT
Progenitor cells maintain self-renewing tissues throughout life by sustaining their capacity for proliferation while suppressing cell cycle exit and terminal differentiation. DNA methylation provides a potential epigenetic mechanism for the cellular memory needed to preserve the somatic progenitor state through repeated cell divisions. DNA methyltransferase 1 (DNMT1) maintains DNA methylation patterns after cellular replication. Although dispensable for embryonic stem cell maintenance, the role for DNMT1 in maintaining the progenitor state in constantly replenished somatic tissues, such as mammalian epidermis, is unclear. Here we show that DNMT1 is essential for epidermal progenitor cell function. DNMT1 protein was found enriched in undifferentiated cells, where it was required to retain proliferative stamina and suppress differentiation. In tissue, DNMT1 depletion led to exit from the progenitor cell compartment, premature differentiation and eventual tissue loss. Genome-wide analysis showed that a significant portion of epidermal differentiation gene promoters were methylated in self-renewing conditions but were subsequently demethylated during differentiation. Furthermore, UHRF1 (refs 9, 10), a component of the DNA methylation machinery that targets DNMT1 to hemi-methylated DNA, is also necessary to suppress premature differentiation and sustain proliferation. In contrast, Gadd45A and B, which promote active DNA demethylation, are required for full epidermal differentiation gene induction. These data demonstrate that proteins involved in the dynamic regulation of DNA methylation patterns are required for progenitor maintenance and self-renewal in mammalian somatic tissue.

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DNMT1 is required to repress differentiation and sustain proliferationa, Venn diagram (left) illustrating overlap between changes identified with DNMT1 loss and calcium-induced differentiation (+Ca2+). Heat map of the 545 genes shared between the two profiles (P-value<1×10-248; Fisher's exact test); red (induced) and green (repressed), log2-based scale. GO analysis (right) (P-values represent a Bonferroni-corrected EASE score). b, QRT-PCR verification of array data. Error bars=s.d., n= 3. c, Cell proliferation; error bars=s.d., n= 2. d, DNMT1 depletion results in G1 cell cycle arrest, flow cytometry profiles; error bars=s.d., n= 2. e, Clonogenic assays of control shRNA and DNMT1i keratinocytes plated at limiting dilution. f, Quantitation of colonies >5 mm2 (n=3/group); error bars=s.d.
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Figure 3: DNMT1 is required to repress differentiation and sustain proliferationa, Venn diagram (left) illustrating overlap between changes identified with DNMT1 loss and calcium-induced differentiation (+Ca2+). Heat map of the 545 genes shared between the two profiles (P-value<1×10-248; Fisher's exact test); red (induced) and green (repressed), log2-based scale. GO analysis (right) (P-values represent a Bonferroni-corrected EASE score). b, QRT-PCR verification of array data. Error bars=s.d., n= 3. c, Cell proliferation; error bars=s.d., n= 2. d, DNMT1 depletion results in G1 cell cycle arrest, flow cytometry profiles; error bars=s.d., n= 2. e, Clonogenic assays of control shRNA and DNMT1i keratinocytes plated at limiting dilution. f, Quantitation of colonies >5 mm2 (n=3/group); error bars=s.d.

Mentions: Global gene expression profiles in DNMT1-deficient cells cultured in growth conditions was next compared to that of control cells induced to differentiate with calcium. 3,343 genes (1,366 induced; 1,977 repressed) changed during calcium-induced differentiation and 797 genes (450 induced; 347 repressed) were altered by DNMT1 loss (Supplementary Tables 1 and 2). Comparing these two gene sets revealed a statistically significant overlap of 545 genes, the vast majority of which were regulated in the same direction (Fig. 3a, b; Supplementary Table 3). Up-regulated genes in this shared set were enriched for differentiation gene ontology (GO) terms, while repressed genes were enriched for proliferation GO terms (Fig. 3a). DNMT1 thus controls an epidermal gene expression program compatible with a role in sustaining proliferation and repressing differentiation.


DNMT1 maintains progenitor function in self-renewing somatic tissue.

Sen GL, Reuter JA, Webster DE, Zhu L, Khavari PA - Nature (2010)

DNMT1 is required to repress differentiation and sustain proliferationa, Venn diagram (left) illustrating overlap between changes identified with DNMT1 loss and calcium-induced differentiation (+Ca2+). Heat map of the 545 genes shared between the two profiles (P-value<1×10-248; Fisher's exact test); red (induced) and green (repressed), log2-based scale. GO analysis (right) (P-values represent a Bonferroni-corrected EASE score). b, QRT-PCR verification of array data. Error bars=s.d., n= 3. c, Cell proliferation; error bars=s.d., n= 2. d, DNMT1 depletion results in G1 cell cycle arrest, flow cytometry profiles; error bars=s.d., n= 2. e, Clonogenic assays of control shRNA and DNMT1i keratinocytes plated at limiting dilution. f, Quantitation of colonies >5 mm2 (n=3/group); error bars=s.d.
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Figure 3: DNMT1 is required to repress differentiation and sustain proliferationa, Venn diagram (left) illustrating overlap between changes identified with DNMT1 loss and calcium-induced differentiation (+Ca2+). Heat map of the 545 genes shared between the two profiles (P-value<1×10-248; Fisher's exact test); red (induced) and green (repressed), log2-based scale. GO analysis (right) (P-values represent a Bonferroni-corrected EASE score). b, QRT-PCR verification of array data. Error bars=s.d., n= 3. c, Cell proliferation; error bars=s.d., n= 2. d, DNMT1 depletion results in G1 cell cycle arrest, flow cytometry profiles; error bars=s.d., n= 2. e, Clonogenic assays of control shRNA and DNMT1i keratinocytes plated at limiting dilution. f, Quantitation of colonies >5 mm2 (n=3/group); error bars=s.d.
Mentions: Global gene expression profiles in DNMT1-deficient cells cultured in growth conditions was next compared to that of control cells induced to differentiate with calcium. 3,343 genes (1,366 induced; 1,977 repressed) changed during calcium-induced differentiation and 797 genes (450 induced; 347 repressed) were altered by DNMT1 loss (Supplementary Tables 1 and 2). Comparing these two gene sets revealed a statistically significant overlap of 545 genes, the vast majority of which were regulated in the same direction (Fig. 3a, b; Supplementary Table 3). Up-regulated genes in this shared set were enriched for differentiation gene ontology (GO) terms, while repressed genes were enriched for proliferation GO terms (Fig. 3a). DNMT1 thus controls an epidermal gene expression program compatible with a role in sustaining proliferation and repressing differentiation.

Bottom Line: DNMT1 protein was found enriched in undifferentiated cells, where it was required to retain proliferative stamina and suppress differentiation.Genome-wide analysis showed that a significant portion of epidermal differentiation gene promoters were methylated in self-renewing conditions but were subsequently demethylated during differentiation.These data demonstrate that proteins involved in the dynamic regulation of DNA methylation patterns are required for progenitor maintenance and self-renewal in mammalian somatic tissue.

View Article: PubMed Central - PubMed

Affiliation: Programs in Epithelial Biology and Cancer Biology and the Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California 94305, USA.

ABSTRACT
Progenitor cells maintain self-renewing tissues throughout life by sustaining their capacity for proliferation while suppressing cell cycle exit and terminal differentiation. DNA methylation provides a potential epigenetic mechanism for the cellular memory needed to preserve the somatic progenitor state through repeated cell divisions. DNA methyltransferase 1 (DNMT1) maintains DNA methylation patterns after cellular replication. Although dispensable for embryonic stem cell maintenance, the role for DNMT1 in maintaining the progenitor state in constantly replenished somatic tissues, such as mammalian epidermis, is unclear. Here we show that DNMT1 is essential for epidermal progenitor cell function. DNMT1 protein was found enriched in undifferentiated cells, where it was required to retain proliferative stamina and suppress differentiation. In tissue, DNMT1 depletion led to exit from the progenitor cell compartment, premature differentiation and eventual tissue loss. Genome-wide analysis showed that a significant portion of epidermal differentiation gene promoters were methylated in self-renewing conditions but were subsequently demethylated during differentiation. Furthermore, UHRF1 (refs 9, 10), a component of the DNA methylation machinery that targets DNMT1 to hemi-methylated DNA, is also necessary to suppress premature differentiation and sustain proliferation. In contrast, Gadd45A and B, which promote active DNA demethylation, are required for full epidermal differentiation gene induction. These data demonstrate that proteins involved in the dynamic regulation of DNA methylation patterns are required for progenitor maintenance and self-renewal in mammalian somatic tissue.

Show MeSH
Related in: MedlinePlus