Xenbase Image ID: 137346
Extended Data Figure 7. cp110 is a direct target of miR-34/449 miRNAsa. A schematic representation of two predicted miR-34/449 binding sites in the mouse cp110 3′UTR and in the luciferase reporter construct that contains cp110 3′UTR. b. Cp110 protein levels at postnatal day 23 are elevated in miR-34/449 TKO tracheal epithelia. c. The expression of Luc-cp110-3′UTR exhibits miR-34b-dependent repression in NIH/3T3 cells. Error bars represent s.e.m., n=3. Paired t-test, * P < 0.05. d. A schematic representation of one predicted miR-34/449 binding site in the frog cp110 3′UTR. A truncated cp110 construct, cp110δ3′UTR, was made to generate a cp110 cDNA without the miR-34/449 target site. e. Real time PCR monitoring cp110 reveals elevated mRNA expression levels of cp110 in miR-34/440 morphant frog skin explants as compared to Ctrl MO injected specimens. f. Timeline of MCC ciliation and recapitulation of ciliation defects in skin explants (animal caps). Representative confocal images from staged whole embryos and skin explants injected with either Ctrl MO or miR-34/449 MOs show the onset of ciliation at stage 26 and fully ciliated skin ectoderm at stage 32 in whole embryos and Ctrl MO injected skin explants. miR-34/449 MOs injected skin explants develop MCC ciliogenesis defects comparable to whole embryo treatment. Cilia: Ac-α-Tub (red), Actin: Phalloidin-488 (green) and nuclei: DAPI (blue). g. Expression of cp110, foxj1 and miR-34/449 RNAs during time course of ciliation in skin explants. Explants at stage 10 represent unciliated MCC precursors, explants at stage 26 represent MCCs at the onset of ciliation, and stage 32 explants represent fully ciliated ectodermal epithelium. cp110 mRNA levels decrease over the time course of ciliation, with the strongest decrease between stage 10 and 26, while foxj1 mRNA levels rapidly increase during this time. miR-34a, -34b and -449c levels strongly increase between stage 10 and stage 26; and only a moderate increase or even decrease can be observed between stage 26 and 32, similar to foxj1 expression levels. Error bars represent s.e.m. n=2, technical replicates on pools of 30 skin explants for each time point.
Image published in: Song R et al. (2014)
Image downloaded from an Open Access article in PubMed Central. Image reproduced on Xenbase with permission of the publisher and the copyright holder.
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