XB-IMG-172713
Xenbase Image ID: 172713
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Fig. 6. KDM3A-mediated demethylation of
H3K9me2 enhances Neurog2 recruitment
at the neurod1 promoter. (A) Western blot
detection of soluble histone H3, H3K9me1,
H3K9me2 and H3K9me3 in control and 2 ng
kdm3a mRNA-injected embryos. (B) Embryos
stained with red-Gal and in situ hybridized with
neurod1 (left panel, stage 18) or tubb2b (right
panel, stage 15). β-gal (200 pg) together with
1 ng kdm3a was injected into one cell at the
two-cell stage. (C) Western blot detection of
chromatin histone H3, H3K9me1, H3K9me2
and H3K9me3 in embryos without injection or
injected with 80 ng cMO or 3A MO. See
Materials and Methods for isolation of
chromatin histones. (D) Anti-H3K9me2 ChIPqPCR
analyses showing that 80 ng KDM3A
MO increased the H3K9me2 marks on the
neurod1 promoter. (E) Anti-H3K9me2 ChIPqPCR
analyses showing that Neurog2 was
unable to reduce the H3K9me2 marks on the
neurod1 promoter when KDM3A was depleted
by injecting 80 ng 3A MO. (F) Anti-Myc ChIPqPCR
analyses showing that ectopic Neurog2
was unable to bind the neurod1 promoter when
KDM3A was depleted (cMO or 3A MO: 80 ng).
ChIP-qPCR results shown in D-F were
combined from two biological repeats. Three
technical replicates were made in each
biological repeat. *P<0.05; **P<0.01;
***P<0.005, ns, not significant (two-tailed
Student’s t-test). (G) A summary of our current
findings that posit the roles for KDM3A in
regulating chromatin states and its
collaboration with Neurog2 to initiate neuronal
precursor differentiation. See details in the
Discussion.
3681 Image published in: Lin H et al. (2017) Copyright © 2017. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License.
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