BTF3 is a potential new substrate of protein kinase CK2.
BTF3, initially discovered as a factor required for transcription inititation of RNA polymerase II, is expressed in two isoforms, termed a and b. BTF3b, the transcriptionally inactive isoform, was identified as an interaction partner of protein kinase CK2 subunit beta employing the interaction trap system for screening ofa HeLa cDNA fusion library. We report here on the interaction between the other isoform, BTF3a, and protein kinase CK2. The complete 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 BTF3a with CK2 subunits alpha, alpha' or beta was detectable by one of three reporters, whereas the CK2beta - BTF3a interaction was activating two reporters. It was also shown that BTF3a is phosphorylated in vitro by the alpha2beta2 holoenzyme, but not by alpha or alpha' alone, indicating the requirement of beta for substrate recognition. Immunoprecipitations of GST-fused BTF3a carried out in vitro resulted in co-precipitation of beta. Similarly, GST-BTF3a, but not GST alone isolated with glutathione agarose beads from buffer containing recombinant CK2 subunits was found complexed with alpha and beta, likely representing alpha2beta2 holoenzyme. The data show a weak, nevertheless specific interaction of protein kinase CK2 via subunit beta with the putative transcription factor BTF3a in vitro and in vivo, and a role of BTF3a as a potential new substrate for CK2.
Grein
S
S
Biochemische Zellphysiologie, Deutsches Krebsforschungszentrum, Heidelberg, Germany.
Pyerin
W
W
D000595:Amino Acid Sequence; D001483:Base Sequence; D047390:Casein Kinase II; D017931:DNA Primers; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013379:Substrate Specificity; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*
D017931:DNA Primers; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C063552:transcription factor BTF3; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases