Cloning of a putative Bombyx mori TFIIB‐related factor (BRF)

Abstract

To identify the protein domains responsible for its conserved and specialized functions, putative TFIIB-Related Factor (BRF) from the silkworm (Bombyx mori) was compared with BRFs from other organisms. The Bombyx BRF coding region was assembled from three separate and overlapping cDNA fragments. Fragments encoding the middle portion and the 3' end were discovered in the Bombyx mori Genome Project "Silkbase" collection through sequence homology with human BRF1, and the fragment encoding the N-terminus was isolated in our laboratory using the 5' RACE method. Southern analysis showed that silkworm BRF is encoded by a single-copy gene. Bombyx BRF contains the following domains that have been noted in all other BRFs, and that are likely, therefore, to provide highly conserved functions: a zinc finger domain, an imperfect repeat, three "BRF Homology" domains, and an acidic domain at the C-terminus. As expected from the evolutionary relationships among insects and mammals, Bombyx BRF is more similar overall to Drosophila BRF (55% identical) than to human BRF1 (42% identical). Detailed examination of individual domains reveals a remarkable exception, however. Domain II of Bombyx BRF is more similar to its human counterpart than to Drosophila Domain II. This result indicates that Domain II has undergone unusual divergence in Drosophila, and suggests a structural basis for Drosophila BRF's unique pattern of interaction with other transcription factors.

Mapping the Protein Interaction Network for TFIIB-Related Factor Brf1 in the RNA Polymerase III Preinitiation Complex

“…Although this Bdp1-binding block contains low sequence homology, secondary-structure analysis indicates consensus ␣-helical secondary structures in this region. Furthermore, this Bdp1-binding block contains the amino acid sequence Gly328-Glu329-Gln330-Glu331-Leu332 (GEXEL), which was previously reported to be a conserved short motif in Brf1c (25). The structural region of Bdp1 that interacts with this sequence block remains to be determined.…”

“…Yeast Brf1c (aa 287 to 596) contains three homologous sequence blocks, I (aa 287 to 304), II (aa 461 to 515), and III (aa 570 to 596) (Fig. 1A), that are conserved in S. cerevisiae, Schizosaccharomyces pombe, Candida albicans, Kluyveromyces lactis, and Homo sapiens (22,25). Brf1c exists mostly as a scaffold that holds together the three TFIIIB subunits (12, 26).…”

Transcription of individual tRNAGly1 genes from within a multigene family is regulated by transcription factor TFIIIB

“…We believe that TBP and not TRF is involved as the component of TFIIIB in RNA pol III transcription in B. mori and the presence of TRF could be exclusive to Drosophila . The recent characterization of the cDNA encoding Brf1 from B. mori [42] has revealed that individual domains of Brf had considerable similarity to the Drosophila counterpart (55%). However, the domain II, which interacts with TBP in most cases but with TRF in Drososphila , was divergent in B. mori .…”