ZFIN ID: ZDB-FIG-161018-29
Olsen et al., 2016 - G9a and ZNF644 Physically Associate to Suppress Progenitor Gene Expression during Neurogenesis. Stem Cell Reports   7(3):454-70 Full text @ Stem Cell Reports
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Antibody:
Fish:
Knockdown Reagents:
Anatomical Terms:
Stage Range: Long-pec to Protruding-mouth
PHENOTYPE:
Fish:
Knockdown Reagents:
Observed In:
Stage Range: Long-pec to Protruding-mouth

Fig. S3

(A) WISH assays monitoring the expression of vsx2 at 48 hpf or otx2 at 72 hpf in lateral views or retinal cross-sections from WT, znf644a morphant or znf644b morphant embryos, as well as morphant embryos rescued by co-injection of the indicated mRNA. Red arrows indicate hindbrain neural cell populations that express vsx2. (B) Frequency at which the indicated embryos exhibited normal or mislocalized expression of vsx2 (top) or otx2 (bottom). (C) WISH assays monitoring the expression of lef1 at 48 hpf (dorsal views) in WT, g9a morphant, znf644a morphant, or znf644b morphant embryos. Red arrows point to lef1-positive midbrain regions, and red arrowheads point to lef1-expressing cells of the anterior hypothalamus. (D) ChIP-PCR assays monitoring the levels of H3K9me2 at the indicated positions near the TSS of vsx2 or ccnd1 genes (as in Figure 3C) at 48 hpf. An anti-H3K9me2 antibody and an isotype control (IgG) antibody were used for ChIP, followed by PCR amplification (35 cycles). Densitometry analysis revealed a ~50% reduction in the levels of H3K9me2 at the ccnd1 promoter, and complete loss of H3K9me2 levels at the vsx2 promoter in g9a morphant embryos. (E) Immunostaining with H3K9me2 antibody demonstrates a global reduction in nuclear expression in the g9a morphant retinas at 48 hpf compared to controls.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
lef1 AB standard conditions Long-pec hypothalamus anterior region ISH
Long-pec midbrain ISH
AB + MO1-ehmt2 standard conditions Long-pec midbrain ISH
AB + MO1-znf644a standard conditions Long-pec midbrain ISH
AB + MO1-znf644b standard conditions Long-pec midbrain ISH
otx2b AB standard conditions Protruding-mouth retina multi fate stem cell ISH
AB + MO1-znf644a standard conditions Protruding-mouth retina multi fate stem cell ISH
AB + MO1-znf644b standard conditions Protruding-mouth retina multi fate stem cell ISH
vsx2 AB standard conditions Long-pec hindbrain neuron ISH
Long-pec retina ISH
AB + MO1-znf644a standard conditions Long-pec hindbrain neuron ISH
Long-pec retina central region ISH
AB + MO1-znf644b standard conditions Long-pec hindbrain neuron ISH
Long-pec retina central region ISH
Antibody Labeling Details
Antibody Assay Fish Conditions Stage Anatomy
Ab34-h3 IHC AB standard conditions Long-pec retina nucleus
IHC AB + MO1-ehmt2 standard conditions Long-pec retina nucleus
Phenotype Details
Fish Conditions Stage Phenotype
AB + MO1-ehmt2 standard conditions Long-pec midbrain lef1 expression decreased amount, abnormal
Long-pec retina nucleus Ab34-h3 labeling decreased amount, abnormal
AB + MO1-znf644a standard conditions Long-pec hindbrain neuron vsx2 expression mislocalised, abnormal
Long-pec hindbrain neuron vsx2 expression spatial pattern, abnormal
Long-pec midbrain lef1 expression decreased amount, abnormal
Long-pec retina central region vsx2 expression mislocalised, abnormal
Protruding-mouth retina multi fate stem cell otx2b expression increased amount, abnormal
Protruding-mouth retina multi fate stem cell otx2b expression spatial pattern, abnormal
AB + MO1-znf644b standard conditions Long-pec hindbrain neuron vsx2 expression mislocalised, abnormal
Long-pec hindbrain neuron vsx2 expression spatial pattern, abnormal
Long-pec midbrain lef1 expression decreased amount, abnormal
Long-pec retina central region vsx2 expression mislocalised, abnormal
Protruding-mouth retina multi fate stem cell otx2b expression increased amount, abnormal
Protruding-mouth retina multi fate stem cell otx2b expression spatial pattern, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Stem Cell Reports for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Stem Cell Reports