Nucleotide sequences of the upstream regions of SDHa, SDHb,
SDHc, and SDHd in H9c2 cells. A, consensus
sequences for transcription factor binding located with DNASIS (Hitachi
Software; Alameda, CA) and MatInspector (Genomatix Software; München,
Germany). Putative NRF-1 and NRF-2 canonical binding sites of 92-100% homology
are underlined (EntrezGene data base ID numbers SDHa 157074,
SDHb 298596, SDHc 289217, SDHd 363061). The
arrows indicate major transcription start sites. Uppercase
letters indicate promoter and intron sequences, and lowercase
letters are exon sequences. a, SDHa 5′-region
within the 180-nt upstream sequence and the NRF-1 sites underlined;
b, SDHb 5′-region within the 180-nt upstream sequence
and NRF-2 sites underlined; the underlined sites at -90 to -77 is a
partial NRF-1 consensus sequence; c, SDHc 5′-region
within the 180-nt upstream sequence, the NRF-2 site is underlined; no
NRF-1 site is found; d, SDHd 5′-region within the 180-nt
upstream sequence, the NRF-2 sites and single NRF-1 site are
underlined. B, ChIP assay for NRF-1 binding to the SDHa,
SDHb, SDHc, and SDHd promoters. H9c2 cells were
transfected with scRNA or siRNA directed at NRF-1 or exposed to 100
μm DCM/CO or medium alone and subjected to ChIP assays using
anti-NRF-1. Input lanes show chromatin PCR product before
immunoprecipitation. Precipitated DNA was analyzed by PCR with specific primer
sets. Data are from one of three experiments.