FIGURE 2.

ELL2 influences IgH secretory mRNA production, histone methylations, and factor associations with the IgH gene. A, The IgH and its mRNA products are diagrammed. Primers used for quantification of mRNA levels are indicated on the respective mRNAs and listed in Table 1. The regions indicated on the gene are: TATA, for transcription start site or TATA box; V:J2, for the V to J fusion region; J4 for the unused joining-region remaining in the intervening sequence; EH for the heavy chain enhancer region also known as Eμ; CH1, for constant heavy chain region 1; CH2, constant heavy chain region 2; sec pA for the secretory specific poly(A) site; sec + 200, the region 200 bp downstream of the secretory poly(A) site; IVS for the intervening sequence between the secretory and membrane specific regions; M1 for membrane-specific exon 1; pAmb3′ for a probe just 3′ of the membrane specific poly(A) site. B, the B cells were transfected with the intact IgH of γ2b on a plasmid and the indicated cDNAs for ELL2, ELL1, or CstF2 or siRNA as described previously. Cells were harvested 48 h after transfection, and the level of IgH secretory and membrane forms of mRNA was determined using quantitative RT-PCR with the probes diagramed in Panel A on the mRNAs. The value for the sec:mb mRNA ratio in plasma cells is shown for comparison. %IP, percentage immunoprecipitated. C and D, a plasma cell lacking its own IgH was co-transfected with an intact IgH gene and siRNA to ell2, a nonsense siRNA, or an ELL2 cDNA lacking the siell2 target sequence. After 48 h, the cells were subjected to chromatin immunoprecipitation with the indicated antibodies. The qPCR signals for the TATA and V-J region of the transfected IgH were averaged from at least three biological replicates. C, ChIP was performed with antibodies to the various H3K methylations. D, ChIP was performed with antibodies to the indicated initiation and elongation factors. ns, normal serum.