CLONING AND CHARACTERIZATION OF THE BETA-SUBUNIT OF HUMAN PROXIMAL SEQUENCE ELEMENT-BINDING TRANSCRIPTION FACTOR AND ITS INVOLVEMENT IN TRANSCRIPTION OF SMALL NUCLEAR-RNA GENES BY RNA-POLYMERASE-II AND RNA-POLYMERASE-III

The proximal sequence element (PSE)-binding transcription factor (PTF) , which binds the PSE of both RNA polymerase II- and RNA polymerase II I-transcribed mammalian small nuclear RNA (snRNA) genes, is essential for their transcription, We previously reported the purification of hu man PTF, a complex of four subunits, and the molecular cloning and cha racterization of PTF gamma and delta subunits. Here we describe the is olation and expression of a cDNA encoding PTF delta, as well as functi onal studies using anti-PTF beta antibodies. Native PTF beta, in eithe r protein fractions or a PTF-Oct-1-DNA complex, can be recognized by p olyclonal antibodies raised against recombinant PTF beta, Immunodeplet ion studies show that PTF beta is required for transcription of both c lasses of snRNA genes in vitro. In addition, immunoprecipitation analy ses demonstrate that substantial and similar molar amounts of TATA-bin ding protein (TBP) and TFIIIB90 fan weakly associate with PTF at low s alt conditions, but this association is dramatically reduced at high s alt concentrations. Along with our previous demonstration of both phys ical interactions between PTF gamma/PTF delta and TBP and the involvem ent of TFIIIB90 in the transcription of class III snRNA genes, these r esults are consistent with the notion that a TBP-containing complex re lated to TFIIIB is required for the transcription of class III snRNA g enes, and acts through weak interaction with the four-subunit PTF.