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Gene regulatory networks in neural cell fate acquisition from genome-wide chromatin association of Geminin and Zic1

View Article: PubMed Central - PubMed

ABSTRACT

Neural cell fate acquisition is mediated by transcription factors expressed in nascent neuroectoderm, including Geminin and members of the Zic transcription factor family. However, regulatory networks through which this occurs are not well defined. Here, we identified Geminin-associated chromatin locations in embryonic stem cells and Geminin- and Zic1-associated locations during neural fate acquisition at a genome-wide level. We determined how Geminin deficiency affected histone acetylation at gene promoters during this process. We integrated these data to demonstrate that Geminin associates with and promotes histone acetylation at neurodevelopmental genes, while Geminin and Zic1 bind a shared gene subset. Geminin- and Zic1-associated genes exhibit embryonic nervous system-enriched expression and encode other regulators of neural development. Both Geminin and Zic1-associated peaks are enriched for Zic1 consensus binding motifs, while Zic1-bound peaks are also enriched for Sox3 motifs, suggesting co-regulatory potential. Accordingly, we found that Geminin and Zic1 could cooperatively activate the expression of several shared targets encoding transcription factors that control neurogenesis, neural plate patterning, and neuronal differentiation. We used these data to construct gene regulatory networks underlying neural fate acquisition. Establishment of this molecular program in nascent neuroectoderm directly links early neural cell fate acquisition with regulatory control of later neurodevelopment.

No MeSH data available.


During neural cell fate acquisition, Gmnn and Zic1 cooperatively promote the expression of Gmnn- and Zic1-associated genes.ES cells that stably over-expressed both Gmnn and Zic1 were established and Gmnn (A) or Zic1 (B) mRNA levels were defined in over-expressing ES cells relative to non-overexpressing cells (cont.). Gmnn and Zic1 protein levels after combined Gmnn + Zic1 over-expression are shown in Figs 5D and 6C. (C,D) Expression levels of four Gmnn- and Zic1-associated genes were evaluated by qRTPCR on days 1–3 of neural fate acquisition, in response to single or combinatorial over-expression of Gmnn and/or Zic1. Gene expression levels on each day of NE fate acquisition are expressed relative to ES = 1.0 and p-values shown (student’s t-test) compare expression with versus without Gmnn and/or Zic1 OE at each day of NE fate acquisition: *** =< 0.001, ** =< 0.01, * =< 0.05, ns = not significant. Error bars represent standard deviation for a representative qPCR performed in triplicate.
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f8: During neural cell fate acquisition, Gmnn and Zic1 cooperatively promote the expression of Gmnn- and Zic1-associated genes.ES cells that stably over-expressed both Gmnn and Zic1 were established and Gmnn (A) or Zic1 (B) mRNA levels were defined in over-expressing ES cells relative to non-overexpressing cells (cont.). Gmnn and Zic1 protein levels after combined Gmnn + Zic1 over-expression are shown in Figs 5D and 6C. (C,D) Expression levels of four Gmnn- and Zic1-associated genes were evaluated by qRTPCR on days 1–3 of neural fate acquisition, in response to single or combinatorial over-expression of Gmnn and/or Zic1. Gene expression levels on each day of NE fate acquisition are expressed relative to ES = 1.0 and p-values shown (student’s t-test) compare expression with versus without Gmnn and/or Zic1 OE at each day of NE fate acquisition: *** =< 0.001, ** =< 0.01, * =< 0.05, ns = not significant. Error bars represent standard deviation for a representative qPCR performed in triplicate.

Mentions: In considering regulatory networks through which Geminin and Zic1 could control neural fate acquisition, associated genes encoding transcription factors with CNS-enriched expression may be of particular interest. Gmnn and Zic1 co-associated transcription factors exhibited CNS-enriched expression specifically at embryonic but not at adult stages (Fig. 7F), while many have known roles in regulating neural development (e.g. Ascl1, Sox11, Irx3, Pou4f1, Hes1, Pax7, Lhx5, Otx1, Myt1l)(Fig. 7G). Gmnn and Zic1 co-association with these genes could reflect a cooperative role in regulating their expression. To test this, we overexpressed either Gmnn or Zic1, or both Gmnn and Zic1 together, during neuroectodermal fate acquisition (Figs 4D, 5C and 8A,B) and assessed levels of four co-associated transcription factors (Fig. 7G) with roles in neurogenesis (Ascl1), regional neural plate patterning (Pax7, Irx3), or neuronal differentiation (Sox11). Neither Gmnn nor Zic1 over-expression singly was sufficient to elevate levels of these genes substantially during neuroectodermal fate acquisition, but combined over-expression of Gmnn and Zic1 cooperatively elevated expression levels of all four genes (Fig. 8C,D). Therefore, association of both Zic1 and Gmnn with subsets of genes encoding transcription factors that control neural development may represent one mechanism for promoting their expression.


Gene regulatory networks in neural cell fate acquisition from genome-wide chromatin association of Geminin and Zic1
During neural cell fate acquisition, Gmnn and Zic1 cooperatively promote the expression of Gmnn- and Zic1-associated genes.ES cells that stably over-expressed both Gmnn and Zic1 were established and Gmnn (A) or Zic1 (B) mRNA levels were defined in over-expressing ES cells relative to non-overexpressing cells (cont.). Gmnn and Zic1 protein levels after combined Gmnn + Zic1 over-expression are shown in Figs 5D and 6C. (C,D) Expression levels of four Gmnn- and Zic1-associated genes were evaluated by qRTPCR on days 1–3 of neural fate acquisition, in response to single or combinatorial over-expression of Gmnn and/or Zic1. Gene expression levels on each day of NE fate acquisition are expressed relative to ES = 1.0 and p-values shown (student’s t-test) compare expression with versus without Gmnn and/or Zic1 OE at each day of NE fate acquisition: *** =< 0.001, ** =< 0.01, * =< 0.05, ns = not significant. Error bars represent standard deviation for a representative qPCR performed in triplicate.
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f8: During neural cell fate acquisition, Gmnn and Zic1 cooperatively promote the expression of Gmnn- and Zic1-associated genes.ES cells that stably over-expressed both Gmnn and Zic1 were established and Gmnn (A) or Zic1 (B) mRNA levels were defined in over-expressing ES cells relative to non-overexpressing cells (cont.). Gmnn and Zic1 protein levels after combined Gmnn + Zic1 over-expression are shown in Figs 5D and 6C. (C,D) Expression levels of four Gmnn- and Zic1-associated genes were evaluated by qRTPCR on days 1–3 of neural fate acquisition, in response to single or combinatorial over-expression of Gmnn and/or Zic1. Gene expression levels on each day of NE fate acquisition are expressed relative to ES = 1.0 and p-values shown (student’s t-test) compare expression with versus without Gmnn and/or Zic1 OE at each day of NE fate acquisition: *** =< 0.001, ** =< 0.01, * =< 0.05, ns = not significant. Error bars represent standard deviation for a representative qPCR performed in triplicate.
Mentions: In considering regulatory networks through which Geminin and Zic1 could control neural fate acquisition, associated genes encoding transcription factors with CNS-enriched expression may be of particular interest. Gmnn and Zic1 co-associated transcription factors exhibited CNS-enriched expression specifically at embryonic but not at adult stages (Fig. 7F), while many have known roles in regulating neural development (e.g. Ascl1, Sox11, Irx3, Pou4f1, Hes1, Pax7, Lhx5, Otx1, Myt1l)(Fig. 7G). Gmnn and Zic1 co-association with these genes could reflect a cooperative role in regulating their expression. To test this, we overexpressed either Gmnn or Zic1, or both Gmnn and Zic1 together, during neuroectodermal fate acquisition (Figs 4D, 5C and 8A,B) and assessed levels of four co-associated transcription factors (Fig. 7G) with roles in neurogenesis (Ascl1), regional neural plate patterning (Pax7, Irx3), or neuronal differentiation (Sox11). Neither Gmnn nor Zic1 over-expression singly was sufficient to elevate levels of these genes substantially during neuroectodermal fate acquisition, but combined over-expression of Gmnn and Zic1 cooperatively elevated expression levels of all four genes (Fig. 8C,D). Therefore, association of both Zic1 and Gmnn with subsets of genes encoding transcription factors that control neural development may represent one mechanism for promoting their expression.

View Article: PubMed Central - PubMed

ABSTRACT

Neural cell fate acquisition is mediated by transcription factors expressed in nascent neuroectoderm, including Geminin and members of the Zic transcription factor family. However, regulatory networks through which this occurs are not well defined. Here, we identified Geminin-associated chromatin locations in embryonic stem cells and Geminin- and Zic1-associated locations during neural fate acquisition at a genome-wide level. We determined how Geminin deficiency affected histone acetylation at gene promoters during this process. We integrated these data to demonstrate that Geminin associates with and promotes histone acetylation at neurodevelopmental genes, while Geminin and Zic1 bind a shared gene subset. Geminin- and Zic1-associated genes exhibit embryonic nervous system-enriched expression and encode other regulators of neural development. Both Geminin and Zic1-associated peaks are enriched for Zic1 consensus binding motifs, while Zic1-bound peaks are also enriched for Sox3 motifs, suggesting co-regulatory potential. Accordingly, we found that Geminin and Zic1 could cooperatively activate the expression of several shared targets encoding transcription factors that control neurogenesis, neural plate patterning, and neuronal differentiation. We used these data to construct gene regulatory networks underlying neural fate acquisition. Establishment of this molecular program in nascent neuroectoderm directly links early neural cell fate acquisition with regulatory control of later neurodevelopment.

No MeSH data available.