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. 2011 Sep 1;2(5):237–242. doi: 10.4161/trns.2.5.17803

Figure 3.

Figure 3

RPRD1A and RPRD1B control the RNAP II phosphorylation state at the LEO1 gene. (A) Association of RPRD1A and RPRD1B with the LEO1 gene. ChIP was performed with anti-RPRD1A or anti-Flag antibodies using soluble protein lysates prepared from untransduced HEK293 cells or HEK293 cells expressing tagged RPRD1B, respectively. Immunoprecipitated DNA was quantified by qPCR with primers recognizing the indicated regions of the LEO1 gene; P, promoter; E, exon; IN, intron; 3U, 3′ UTR; LE, last exon. (B) Overexpression of RPRD1A or RPRD1B inhibits RNAP II phosphorylation at the LEO1 promoter. ChIP experiments using 3E8 and 4E12 antibodies specific for S5P and S7P and N-20 antibody that recognizes all RNAP II phosphoisoforms were performed in HEK293 cells overexpressing the indicated genes. Immunoprecipitated DNA was quantified by qPCR with primers recognizing the LEO1 promoter region. Promoter occupancies for S5P and S7P were normalized to that of total RNAP II detected by the N-20 antibody and represent averages from a minimum of two biological replicates. *, statistically significant (p < 0.05) compared with overexpressed GFP.