CTCF does not repress MYC expression and is required for normal levels of MYC mRNA.

<p>Real time RT-PCR analysis of the wildtype and recombinant MYC alleles residing on human chromosome 8 in B78 cells. (A) The steady-state level of human and mouse MYC RNA normalized to the endogenous mouse GAPDH mRNA. Left panel shows the relative expression of human MYC as a percentage of the expression detected in cells carrying the wildtype (wt) MYC allele. This analysis was performed multiple times for each B78 cell line containing a human MYC allele (wt, n = 7; ΔLg, n = 8; ΔCTCF-N, n = 9; ΔCTCF-NA, n = 6). Each clone was independently analyzed at least twice, and each PCR reaction was performed in duplicate. The bars indicate the mean value (±SEM) for all clones with the specific type of mutation. The right panel shows the expression of the endogenous murine MYC gene relative to the GAPDH gene. Mouse MYC gene expression is expressed as a percentage of the expression that is detected in cells carrying the human chromosome 8 with the wildtype allele. The expression in each of the mutant cells lines varies less than two-fold, indicating unaltered expression of the endogenous mouse MYC gene. (B) Expression of wildtype and mutant human MYC genes in starved (0 Hr) and serum-induced cells (1 Hr, 3 Hr, 9 Hr). RNA was harvested prior to serum induction (0 Hr), and 1 hour, 3 hours, and 9 hours post serum induction and analyzed by real-time RT-PCR. MYC mRNA levels were normalized to murine GAPDH mRNA levels. MYC expression in serum-starved cells was set to 1, and RNA levels at 1, 3 and 9 Hr are expressed as fold-induction over 0 Hr timepoint.