BTF3 is a potential new substrate of protein kinase CK2

BTF3, initially discovered as a factor required for transcription inititati on of RNA polymerase fl, is expressed in two isoforms, termed a and b. BTF3 b, the transcriptionally inactive isoform, was identified as an interaction partner of protein kinase CK2 subunit beta employing the interaction trap system for screening of a HeLa cDNA fusion library. We report here on the i nteraction between the other isoform, BTF3a, and protein kinase CK2. The co mplete cDNA of BTF3a was cloned by RT-PCR and used for analysis in the two- hybrid system with a three-reporter yeast strain. Interaction of BTF3 a wit h CK2 subunits alpha, alpha' or beta was detectable by one of three reporte rs, whereas the CK2 beta - BTF3a interaction was activating two reporters. It was also shown that BTF3a is phosphorylated in vitro by the alpha(2)beta (2) holoenzyme, but not by alpha or alpha' alone, indicating the requiremen t of beta for substrate recognition. Immunoprecipitations of GST-fused BTF3 a carried out in vitro resulted in co-precipitation of beta. Similarly, GST -BTF3a, but not GST alone isolated with glutathione agarose beads from buff er containing recombinant CK2 subunits was found complexed with alpha and b eta, likely representing alpha(2)beta(2) holoenzyme. The data show a weak, nevertheless specific interaction of protein kinase CK2 via subunit beta wi th the putative transcription factor BTF3a in vitro and in vivo, and a role of BTF3a as a potential new substrate for CK2.