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. 2018 Dec 27;47(4):e23. doi: 10.1093/nar/gky1286

Figure 7.

Figure 7.

Targeting Sa-dCas9/LSD1 to PDX1 Area IV is sufficient to decrease PDX1 and disrupt PDX1-target gene expression. (A) UCSC genome browser screenshot of the PDX1 locus in the rat genome (rn5). Highlighted areas demark Area I-IV and gRNAArea IV targeting within Area IV. (B) Schematic of pMAGIC-derived adenoviruses expressing Sa-dCas9/LSD1. ChIP-qPCR analysis of (C) HA epitope tag and (D) H3K27ac enrichment using primers to a control region (Chr15) or Area IV in INS1 832/13 cells. Data are shown as a percent of input. (E) qRT-PCR analysis of PDX1 expression levels in INS1 832/13 cells. (F) Immunoblot analysis of INS1 832/13 cell lysates from 3 independent experiments. (G) qRT-PCR gene expression analysis in INS1 832/13 cells for genes directly regulated by PDX1. qRT-PCR analysis of (H) PDX1 and (I) the PDX1 target gene SLC2A2 (GLUT2) in intact primary rat islets. For all experiments, cells were harvested 48 h after transduction with either (–) gRNA control (white) or gRNAArea IV (black) purified adenoviruses depicted in panel B. Data represent mean ± S.E.M. of 3 (E, G) or 4 (C, D, H, I) independent experiments. #P < 0.05 compared to (–) gRNA control adenovirus.