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Fig. 1. Id3 is a Myc target in the neural crest. (A) In situ hybridization showing
the expression of Xenopus Id3 at stage 14 (left) and stage 19
(right). Strong expression is seen in both the neural crest-forming region of
the neural plate border, and in the transverse neural fold, at open neural
plate stages. (B) Following MO-mediated depletion of Myc protein, expression
of Id3 is greatly diminished in neural crest-forming regions on the
injected side of the embryo (arrowheads). (C,D) Id3 rescue of Myc-depleted
embryos. (C) In situ hybridization showing loss of Slug expression on
Myc-depleted side of the embryo (arrowhead). (D) Expression of Slug
is substantially rescued when mRNA encoding Id3 is subsequently injected into
the Myc depleted side of the embryo. Red staining denotes expression of the
lineage tracer ß-galactosidase. Arrowheads indicate the injected side of
the embryo. (E) Western blot analysis demonstrating that Myc and Id3 are
co-expressed in the human glioma cell line T98G. Neither protein is expressed
in adult human brain at detectable levels. (F) ChIP assay demonstrating a
direct interaction between Myc and the Id3 promoter. Chromatin was extracted
from logarithmically growing T98G cells and protein-DNA complexes were
immunoprecipitated with a polyclonal antibody against Myc
(
-Myc) or normal rabbit immunoglobulins (NRIg).
Precipitated DNA was analyzed by PCR with primers from the Id3 and,
as negative control, the OLR1 promoters. PCR was also performed in
the absence of DNA (H2O) and from 0.01% of the total protein-DNA
complexes used in each immunoprecipitation (input).
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