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Figure 4.
CASZ1 Directly Activates Egfl7 Transcription
(A) Illustration of cardiovascular-enriched region dissected from X. tropicalis for ChIP.
(B) Genomic structure of the Xenopus Egfl7 locus, denoting a CASZ1 ChIP fragment. White boxes: exons; shaded boxes: miR-126 in intron 7 and intronic region potentially containing CASZ1 element (�4 kb).
(C�J) In situ analysis of Egfl7 of stage 29�39 control and CASZ1-depleted embryos (lateral view with anterior to the left). Note the downregulation of Egfl7 in the vvn and isv of CASZ1-depleted embryos.
(K) Relative mRNA expression of Egfl7, miR-126, and Flk1 after infection of HUVECs with shCasz1. mRNA levels relative to Rps29 � SEM. ∗∗∗p < 0.001; NS: not significant.
(L) Schematic demarcating Egfl7 genomic DNA regions (in bp) tested for transcriptional activation. E1 (−55 to 1,614) within intron 3 but not E2 (1,773�3,840) resulted in increased luciferase (luc) activity.
(M) Egfl7 genomic region E1 in the presence or absence of Casz1. Bars represent fold increase in activity relative to control � SEM. Experiments were repeated twice on independent batches of embryos; ∗∗p < 0.01.
(N) Identification of a 90 bp region endogenously bound by CASZ1 located within a nonoverlapping region of the E1.2 (113�227) PCR amplicon.
See also Figure S3 and Table S1. |
