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Fig. 8. Tbx20 directly binds and represses Isl1 in myocardium. (A-D) Isl1 is
expressed throughout myocardium in Tbx20 mutants. (E) ChIP analysis
revealed Tbx20 recruitment to region of Isl1 promoter with conserved T-box
site (lane 1, primer P-842, P-581). ChIP analysis with primers against an
unrelated promoter region revealed no Tbx20 recruitment (lane 2) (see
Materials an methods for primers). No recruitment was found with beads, IgG or
H2O. (F) Co-transfection of Tbx20 expression vector (500 ng) with
Isl1 promoter-luciferase reporter (100 ng) demonstrated repression of
Isl1 by Tbx20. Repression was abrogated by mutation of the consensus
T-box element within the Isl1 promoter.
*P<0.05, paired t-test. (G-J) The nebulette
promoter, expressed exclusively in myocardium during early embryogenesis
(J.C., unpublished) was used to drive expression of Isl1 in transient
transgenic mice. Whole-mount in situ hybridization (G,H) revealed expression
of Isl1 throughout myocardium in transgenic mice (H). Hearts of transgenics
appeared normal at E10.5. The left and right images in A-D,G,H show right and
left side views. Corresponding sections are shown progressively from anterior
to posterior (C,D; I,J). Arrows indicate the corresponding heart regions in
wild type and mutant embryos.
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