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. 2018 Jun 1;32(11-12):781–793. doi: 10.1101/gad.315127.118

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© 2018 Cai et al.; Published by Cold Spring Harbor Laboratory Press

This article is distributed exclusively by Cold Spring Harbor Laboratory Press for the first six months after the full-issue publication date (see http://genesdev.cshlp.org/site/misc/terms.xhtml). After six months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.

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Figure 4. — c-Myc is an essential transcriptional target of YAP and a critical effector of cystogenesis in Pkd1 mutant kidneys. (A) Up-regulation of c-Myc in Sav1 mutant (AhCre;Sav1^flox/flox) kidneys 3 mo after P8–P11 β-NF injections. Data are mean ± SD. n = 3. (*) P < 0.01, t-test. (B) Schematic representation of the mouse c-Myc locus showing ∼5 kb of promoter, exons (black boxes), and introns. Red bars indicate potential YAP/TEAD-binding sites. (C) ChIP assay in TCMK-1 cells showing YAP binding to site 2 in the c-Myc promoter and site 6 in the first intron of c-Myc. Data are mean ± SD. n = 3. (*) P < 0.01, t-test. (D) Luciferase reporter assay in HEK293 cells for enhancers containing either wild-type or mutated TEAD-binding sites in the c-Myc locus. Data are mean ± SD. n = 3. (*) P < 0.01, t-test. (E,F) H&E (E) and Ki67 (F) staining in control (Sav1^flox/flox), Sav1 mutant (AhCre; Sav1^flox/flox), Sav1;c-Myc double-mutant (AhCre;Sav1^flox/flox;c-Myc^flox/flox), and c-Myc mutant (AhCre;c- Myc^flox/flox) kidneys 3 mo after P8–P11 β-NF injections. Arrows mark representative kidney dilations (E) and proliferating cells (F), while the arrowhead indicates a glomerular dilation (E). Bar, 100 µm. (G) Quantification of Ki67-positive cells in control, Sav1 mutant, Sav1;Yap double-mutant (AhCre;Sav1^flox/flox;Yap^flox/flox), Yap mutant (AhCre; Yap^flox/flox), Sav1;c-Myc double-mutant, and c-Myc mutant kidneys (as in Fig. 4F; Supplemental Fig. S3A). Data are mean ± SD. n = 3. (*) P < 0.01, t-test. (H) H&E staining of the kidneys in 1- and 3-mo-old Pkd1 mutant and Pkd1;c-Myc double-mutant (AhCre;Pkd1^flox/flox;c-Myc^flox/flox) mice. Bar, 1 mm. (I) The ratio of kidney weight (KW) over body weight (BW) in control (1 mo, n = 5; 3 mo, n = 4), Pkd1 mutant (1 mo, n = 9; 3 mo, n = 5), Pkd1;c-Myc double-mutant (1 mo, n = 7; 3 mo, n = 10), and c-Myc mutant (AhCre; c-Myc^flox/flox) (1 mo, n = 5; 3 mo, n = 5) mice. Data are mean ± SD. (*) P < 0.05, t-test. (J) The cystic index in Pkd1 mutant (1 mo, n = 9; 3 mo, n = 5) and Pkd1;c-Myc double-mutant (1 mo, n = 7; 3 mo, n = 9) kidneys. Data are mean ± SD. (*) P < 0.05, t-test.