Distinct and separable roles for EZH2 in neurogenic astroglia
- University of California, San Francisco, United States
- Veterans Affairs Medical Center, University of California, San Francisco, USA
- University of California, San Francisco, USA
Figures
Figure 1 with 1 supplement
EZH2 is specifically expressed in the neurogenic SVZ lineage.
(A) Schematic of sagittal brain section depicting the RMS (red) that connects the SVZ to the OB. Lines indicate locations of Figure 1B,C. (B and C) Schematic of coronal sections through the SVZ (B) …
Figure 1—figure supplement 1
EZH2 Expression is maintained in the SVZ and SGZ but not the cortex and striatum postnatally.
(A) Schematic of coronal brain section depicting the location of images in (B). Ctx = Cortex, Str = Striatum. (B) IHC analysis of the SVZ, cortex, and striatum staining for DAPI, EZH2 and Vimentin …
Figure 2 with 3 supplements
Conditional deletion of Ezh2 in SVZ NSCs both in vivo and in vitro inhibits neurogenesis.
(A) IHC for the neuroblast marker DCX (green) in P21 OB coronal sections comparing Control to hGFAP-Cre;Ezh2F/F slices. (DAPI; blue). (B and C) IHC for NeuN+ EdU+ cells in the granule cell layer of …
Figure 2—figure supplement 1
Loss of EZH2 and H3K27me3 upon conditional deletion of Ezh2 in vivo and in vitro.
(A) IHC co-localization of EZH2 (green) and H3K27me3 (red) merged with DAPI (blue) in P21 control and hGFAP-cre;Ezh2F/F animals at the SVZ, RMS, and OB. Scale bars, 50µM. (B) ICC co-localization of …
Figure 2—figure supplement 2
Morphology of neurogenic brain regions in Control and hGFAP-cre;Ezh2F/F animals.
Hematoxylin and eosin (H&E) or DAPI staining of brain slices from the cerebellum, hippocampus, and olfactory bulb in P21 control and hGFAP-cre;Ezh2F/F animals.
Figure 2—figure supplement 3
Conditional deletion of Ezh2 in SVZ NSCs does not cause defects in type C or B cells in the SVZ.
(A) IHC for the type C/A marker DLX2 (green) or type A neuroblast marker DCX (red) in P21 OB coronal sections comparing Control to hGFAP-Cre;Ezh2F/F slices. Type C cells are DLX2+ DCX−. (B) …
Figure 3 with 1 supplement
Deletion of the Ink4a/Arf locus rescues proliferation but not neurogenesis in Ezh2Δ/Δ SVZ NSCs.
(A and B) ChIP-qPCR analysis of HoxA2b (positive control), Gapdh (negative control), Arf, Ink4a, and Olig2 genomic loci pulled down by either IgG (non-specific control) or antibodies to EZH2 (A) and …
Figure 3—figure supplement 1
Schematic of the location of ChIP primers to Ink4a/Arf and Olig2 used in Figure 3A,B.
https://doi.org/10.7554/eLife.02439.010
Figure 4 with 5 supplements
Aberrant expression of OLIG2 in Ezh2Δ/Δ;Ink4a/Arf−/− SVZ NSCs inhibits neuronal differentiation.
(A) ChIP-seq analysis of proliferating SVZ NSCs in vitro with an H3K4me3 or H3K27me3 antibody at the Hoxa2, Gapdh, and Olig2 genomic loci. (B) ChIP-qPCR analysis of FACS-sorted Tuj1+ and GFAP/Nestin …
Figure 4—figure supplement 1
FACS analysis of SVZ NSCs after 0 or 4 days of differentiation.
(A) Scatter plot of FACS performed on undifferentiated SVZ NSCs using GFAP and Nestin antibodies. (B) Scatter plot of FACS performed on SVZ NSCs after 4 days in differentiation conditions using a …
Figure 4—figure supplement 2
Aberrant expression of OLIG2 in Ezh2Δ/Δ;Ink4a/Arf−/− SVZ NSCs inhibits neuronal differentiation.
(A) Images of differentiating rtTA-expressing SVZ NSCs infected with a LV expressing GFP and encoding tetO-Olig2. Arrows point to GFP+ cells expressing OLIG2 upon addition of Dox in top panel. (B) …
Figure 4—figure supplement 3
Identification of transcriptional modules that require EZH2 for proper downregulation during differentiation.
To identify other potential targets of Ezh2-dependent gene repression besides Olig2 that are related to SVZ NSC neurogenesis, we analyzed by expression microarray the transcriptional profiles of Ezh2…
Figure 4—figure supplement 4
Ezh2-dependent genes during early differentiation are enriched for homeobox-containing neuronal transcriptional regulators and H3K27me3.
(A) Selected functional categories of the combined genes identified from the saddlebrown, white, and darkgreen modules that are significantly enriched after DAVID analysis (Huang da et al., 2009). (B…
Figure 4—figure supplement 5
List of genes identified in the saddlebrown, white, and darkgreen modules.
Interestingly, many of the genes that exhibited Ezh2-dependent transcriptional repression encode for homeodomain transcription factors involved in the production of neuronal subtypes that are …
Figure 5
EZH2 expression in the infant human subventricular zone decreases post-birth.
(A) Schematic illustrating the location of the 30 µm thick sections of the anterior subventricular zone stained in (C) and (D). RMS = rostral migratory stream; LV = lateral ventricle. (B) …
Figure 6
Model of EZH2 function in maintaining the neurogenic potential of SVZ astroglia.
EZH2 represses multiple targets to promote distinct and separable aspects of adult SVZ neurogenesis.
Additional files
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Supplementary file 1
Quantity and dilution ratios of the primary antibodies used in this study.
- https://doi.org/10.7554/eLife.02439.019
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Supplementary file 2
Primers used for ChIP-qPCR experiments in this study.
- https://doi.org/10.7554/eLife.02439.020
