Figure 4: Transcriptional control of
c-Myc by c-Jun~Fra-2 in the
liver
A. Immunoblot analyses of c-Myc in liver extracts from
c-Jun~Fra-2hep mice (non-tumoral and
tumors) and controls. B. qRT-PCR quantification of c-Myc target
genes c-Jun~Fra-2hep livers at 2
months, p<0.05 for each mRNA. C. Schematic of the
murine c-myc gene indicating the putative AP-1 binding site
elements (TGACTCA) in the promoter and the 3´enhancer (WRE). D.Fra-2 ChIP-qPCR for the c-Myc 3´enhancer (WRE) and promoter in
livers from c-Jun~Fra-2hep mutants and
controls at 2 months of transgene expression (off Dox at weaning). An
AP-1 binding sequence from the Dusp1 promoter and an intergenic
area are included as positive and negative controls, respectively.E. Flag and IgG ChIP-qPCR for the c-Myc 3´enhancer (WRE)
and an intergenic area in livers from
c-Jun~Fra-2hep and
Fra-2hep mutants and controls at 2 months. F.Flag ChIP-qPCR for the c-Myc 3´enhancer (WRE) and Dusp1promoter in livers from
c-Jun~Fra-2hep,
Fra-2hep mutants and controls at 2 months. G.Functional validation in the murine AML12 liver cell line by transient
transfection of Fra-2 or c-Jun~Fra-2 expression vectors,
followed by c-myc qRT-PCR (left) or luminescence measurement of a
co-transfected c-Myc 3´enhancer luciferase reporter (right). H.c-jun , fra-2 and c-myc qRT-PCR in livers from HBV
transgenic mice (HBVtg), lacking c-junexpression (c-JunΔli) in the liver compared to
c-Jun-proficient HBVtg littermates at 3 months of age.I. c-Myc immunoblot in liver extracts from
c-JunΔli HBV transgenic mice compared to
HBVtg c-Jun-proficient littermates over time. 2
samples from mice not carrying the HBV transgene are included. Tubulin
and Actin are used to control immunoblots loading. Bars = means ±SEM,
n≥3. * p<0.05, ** p<0.01, *** p<0.001.