Fig. 4: Characterization of ccndx enhancer regions and negative regulation of ccndx expression by Notch signaling. | Nature Communications

Fig. 4: Characterization of ccndx enhancer regions and negative regulation of ccndx expression by Notch signaling.

From: Stem and progenitor cell proliferation are independently regulated by cell type-specific cyclinD genes

Fig. 4

A UCSC zebrafish genome browser track showing two regions of conservation with the goldfish genome upstream of ccndx exon 1. The black bar indicates the 4.5 kb region cloned. B, C DAPI stained 32 hpf zebrafish larvae showing H2B-EGFP (cyan) expression in the hindbrain, spinal cord and migrating lateral line primordium driven by the ccndx upstream region with a higher magnification image of the primordium (C). Scale bars = 10 µm and 200 µm, respectively. D, E DAPI stained 5 dpf zebrafish showing H2B-EGFP (cyan) expression in the spinal cord and neuromasts driven by the ccndx upstream region with higher magnification view of a neuromast from a different larva (E). Scale bars =10 µm and 200 µm, respectively. ccndx HCR (yellow) in 5 dpf sibling (F) or atoh1a−/− neuromasts (G). sqEt4:EGFP expression (cyan) shows lack of hair cells in atoh1a−/− larvae. ccndx is expressed in atoh1a−/− neuromasts and shows a broader and more central expression pattern. Scale bar = 10 µm. HCR for ccndx (yellow) and atoh1a (magenta) in heat shocked sibling (H) and hs:atoh1a larvae (I). Scale bar = 10 µm. J ccndx HCR (yellow) in zebrafish expressing the tp1bglobin:egfp Notch reporter (cyan) showing the lack of co-localization. Scale bar = 10 µm. HCR for ccndx (yellow) and egfp (magenta) in ccndx:NLS-d2GFP (cyan) transgenic zebrafish treated with DMSO (K) or LY411575 (L) for 6 hrs. LY411575 treatment induces an increase in ccndx and egfp expression, especially in the central region. Scale bar = 10 µm.Maximum projection still images from time lapses of ccndx:NLS-d2EGFP transgenic zebrafish which were treated with DMSO (M) or LY411575 (N) immediately after neomycin treatment. There is an increase in the number of NLS-d2EGFP+ cells around 10 h after neomycin which decreases in DMSO treated fish but continues to increase with Notch inhibition. Scale bar = 10 µm. HCR for ccndx (yellow) in 5 dpf neuromasts in DMSO treated sibling (O) or atoh1a−/− (P) or LY411575 treated sibling (Q) or atoh1a−/− (R). Scale bar = 10 µm.

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