hTFIIIB50 is specifically required for RNA polymerase III transcription
of genes with upstream promoter elements. (Upper)
Transcription of the 7SK gene. (Lower) Transcription of
the human tRNAMET gene. System A was comprised of 1.5 μl
cEDF 1 M fraction, 40 ng rhTBP, and 24 ng rhTFIIIB150. System B
consisted of 1.5 μl cEDF 1 M fraction and 5 μl B-EDF 0.3 fraction
(unpublished observations). The following fractions were added: lane 1,
none; lanes 2 and 3, 5 and 10 μl cEDF 0.2, respectively; and lanes 4
and 5, 5 and 10 μl of flow-through (FT), respectively, of an
anti-hTFIIIB50 antibody column onto which a cEDF 0.2 fraction had been
loaded. Purification of the cEDF fractions and antibody depletion
procedures are described in Materials and Methods.