FIGURE 6.

ATF4 induced CSE transcription via direct binding to a cis regulatory intronic site. A and B, luciferase activity resulting from HEK cells transfected with constructs as indicated together with (empty vector control plasmid) pCDNA3.1 or overexpressed ATF4. RLU, relative light units. n = 4. *, p < 0.05; ****, p < 0.0001. n/s, not significant. C, schematic representation of alignment of putative ATF4-binding sites within intron 1 of mouse and human CSE gene loci. The genomic fragments contained in each luciferase construct are indicated. D and E, formaldehyde cross-linked chromatin prepared from HEK cells transfected with pCDNA3.1 or ATF4 incubated with normal rabbit IgG (negative control), anti-acetyl-histone H3 (positive control), or anti-ATF4 as indicated. Aliquots of chromatin before immunoprecipitation served as a positive control (input). Purified DNA was analyzed using primers specific for site A or site B as indicated in panel C or exon 3 of Human RLP30. The results presented are representative of three separate experiments.