Supplementary Table 1. TaqMan primer/probes used for real-time PCR analysis

qRT-PCR analysis / Sequence
Endogenous P1 / FP1: GTGTTCCGCGTGATTGAAGAC
RP2: CCCCAGAGAAAGAAGAGGAGTTATAA
Probe: CCCTCGTCCAAGAATGCAAAGCACAT
Endogenous P1+P2 / FP: GTTGGCCCCCGTTGCT
RP: CAGCTTATAATGGATGTACTTCATCACT
Probe: TGGCGCACGCTGGGAGAACAG
Reporter P1 / FP: AAATAGCTGGATTATAACTTCTCTTCTTTCTC
RP: TTGGCGTCTTCCATTTTACCA
Probe: TTTTCCTCTAAGCTTGGCATTCCGGTACTG
Reporter P1+P2 / FP: GGTGTTGGGCGCGTTATT
RP: CTACGGTAGGCTGCGAAATGT
Probe: CAGTTGCGCCCGCGAACGAC
ChIP assays / Sequence
bcl-2 Exon 2 / FP: GGTGGTGGAGGAGCTCTTCA
RP: TTGACGCTCTCCACACACATG
Probe: CCCACCGAACTCAAAGAAGGCCACA
bcl-2 Exon 3 / FP: GATGCTGTCTTAGTTTGCGACTGT
RP: CTGGTAGTGGGAAGCCAAGCT
Probe: CCACCTTAACCCAATCCATGCAAGTCTACA
Reporter P2 / FP: AAATAGCTGGATTATAACTTCTCTTCTTTCTC
RP: TTGGCGTCTTCCATTTTACCA
Probe: TTTTCCTCTAAGCTTGGCATTCCGGTACTG
Murine HS12 / FP: CAGTCCCAGTGGCCTATGC
RP: TCTCACTTTCTGAGGTTGTTTCCA
Probe: CCCCATCCCCAAGGCTGGTCA
Human HS12 / FP: GTGGAGAATCGTGCAAGCTATTT
RP: GTTGTTTGTTCCACACCGAAAA
Probe: TTTCTTGCCCTCTGAGGCTGTTTCCA
Human HS3 / FP: GAAGCCTCTTGTACCCCATCAC
RP: GGCCTGGGCTGTGTTCAC
Probe: CCCACAGGGCTATTTTGGATATTTCTGCA
Human HS4 / FP: AGATGGCGATTTGCATTGG
RP: GAATAGTCAGGAATCCTGCAAACC
Probe: AAGGCTGGCACCCAGGCAGCT

FP1: Forward Primer

RP2: Reverse Primer

SUPPLEMENTARY FIGURE LEGENDS

Supplementary Figure 1. Binding of histone H3 acetylated at lysines 9 and 14 and histone H3 dimethylated at lysine 4 is observed at the endogenous IgH enhancer region in DHL-4 cells.

A.Diagram of the enhancers located downstream of the human IgH C1 and C2 genes.

B.Quantitative ChIP assay of modified histone H3 bound to the human IgH enhancers downstream of C2 in DHL-4 cells.

Supplementary Figure 2. Deregulated bcl-2 promoter activity and usage by the IgH 3’ enhancer regions.

A.Diagram of bcl-2 promoter constructs with different regions of the murine IgH enhancers.

B.Bcl-2 promoter activity in the reporter constructs containing different regions of the IgH enhancers. The constructs are named with the region of the IgH enhancer that is present with the bcl-2 promoter-luciferase gene (pHS12 contains the bcl-2 promoter region with IgH enhancer region HS12, etc). The reporter gene constructs were stably transfected into DHL-4 cells. The luciferase transcripts derived from the bcl-2 promoters in pooled transfectants were determined by qRT-PCR using the primer-probe set located in the luciferase sequence. The transcripts in each stable cell line were normalized to GAPDH expression and transgene copy number and represented as fold increase compared to the transcripts in the enhancerless reporter construct (pHS0).

C.Bcl-2 promoter usage in the different episomal constructs. The P1 and P1+P2 transcripts were determined by qRT-PCR as described for Figure 3C.

Supplementary Figure 3. Expression of IB-SR suppresses NF-B activity in DHL-4 cells.

A reporter construct with 5 copies of the NF-B binding site linked to a TATA element was cotransfected with the IB-SR or control expression vector. 24 hr after transfection, the luciferase activity was determined and normalized to protein content in each sample. The relative luciferase activity is shown as the percentage of luciferase activity in the cells transfected with the control expression vector.