Abstract
Apart from its well-established role in the initiation of transcription, the general transcription factor TFIIB has been implicated in the termination step as well. The ubiquity of TFIIB involvement in termination as well as mechanistic details of its termination function, however, remain largely unexplored. Using GRO-seq analyses, we compared the terminator readthrough phenotype in the sua7-1 mutant (TFIIB(sua7-1)) and the isogenic wild type (TFIIB(WT)) strains. Approximately 74% of genes analyzed exhibited a 2-3-fold increase in readthrough of the poly(A)-termination signal in the TFIIB(sua7-1) mutant compared to TFIIB(WT) cells. To understand the mechanistic basis of TFIIB's role in termination, we performed the mass spectrometry of TFIIB-affinity purified from chromatin and soluble cellular fractions-from TFIIB(sua7-1) and TFIIB(WT) cells. TFIIB purified from the chromatin fraction of TFIIB(WT) cells exhibited significant enrichment of CF1A and Rat1 termination complexes. There was, however, a drastic decrease in TFIIB interaction with CF1A and Rat1 complexes in the TFIIB(sua7-1) mutant. ChIP assays revealed about a 90% decline in the recruitment of termination factors in the TFIIB(sua7-1) mutant compared to wild type cells. The overall conclusion of these results is that TFIIB affects the termination of transcription on a genome-wide scale, and the TFIIB-termination factor interaction plays a crucial role in the process.