Fig. 6
REG1CP tethers FANCJ to the REG3A promoter. a the −2325/−2509 fragment of the REG3A gene was hypersensitive to DNase I. b Deletion (Δ) of the −2325/−2509 fragment abolished luciferase reporter activity of pGL3-basic based REG3A promoter constructs. c FANCJ bound to the -2325/-2509 fragment of REG3A promoter as shown in ChIP assays. d The association between REG1CP and the REG3A gene was required for binding of FANCJ to the REG3A promoter. Relative abundance of REG3A promoter associated with FANCJ was measured using ChIP assays. e Silencing of REG1CP and FANCJ reduced sensitivity of the −2325/−2509 fragment of REG3A promoter to a single-stranded DNA-specific DNase (ssDNase). f REG1CP overexpression did not affect the increase in H3K27me3 enrichment to REG3A promoter caused by FANCJ silencing as measured using ChIP assays. g REG1CP silencing diminished reduction in H3K27me3 enrichment to REG3A promoter caused by overexpression of FANCJ. Relative enrichment of H3K27me3 to REG3A promoter was measured using ChIP assays in LIM1215-shREG1CP and HT-29-shREG1CP cells with or without overexpression of FANCJ in the presence or absence of doxycycline (Dox) to induce REG1CP silencing. (h) REG1CP overexpression did not affect the decrease in H3K4me3 enrichment to REG3A promoter caused by FANCJ silencing measured using ChIP assays. i REG1CP silencing diminished the increase in H3K4me3 enrichment to REG3A promoter caused by FANCJ overexpression. Relative enrichment of H3K4me3 to REG3A promoter was measured using ChIP assays in LIM1215-shREG1CP and HT-29-shREG1CP cells with or without overexpression of FANCJ in the presence or absence of doxycycline (Dox) to induce REG1CP silencing. j REG1CP overexpression increased the enrichment of FANCJ to the REG3A promoter. Relative abundance of REG3A promoter associated with FANCJ was measured using ChIP assays. k, l FANCJ silencing reduced (k), whereas its overexpression increased (l) RNA polymerase II (Pol II) enrichment to REG3A promoter measured using ChIP assays. n = 3 independent experiments. Data are presented as Mean ± SEM (b, c, d, f–l) or representatives (a, e). Statistical significance was calculated using a two-tailed t-test.
