Biomedical subjects
Toshio Fukasawa
Publications and source records attributed to Toshio Fukasawa.
A unified nomenclature for protein subunits of mediator complexes linking transcriptional regulators to RNA polymerase II.
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Carboxy-terminal region of the yeast heat shock factor contains two domains that make transcription independent of the TFIIH protein kinase.
BACKGROUND: Phosphorylation of the carboxy-terminal domain (CTD) of the largest subunit of RNA polymerase II is implicated in transition from initiation to elongation in the transcription cycle. In yeast cells, Kin28, a subunit of the general transcription factor TFIIH, is responsible for the CTD phosphorylation. Although Kin28 is indispensable for transcription of many genes, its requirement is bypassed in certain genes such as SSA4 or CUP1, whose transcription is activated by the heat shock factor Hsf1. RESULTS: We show that C-terminal region of Hsf1, which consists of an activation domain AR2 and a regulatory domain CTM, mediates the Kin28-independent transcription. The AR2 domain, when fused to the DNA-binding domain of Gal4 and recruited to the GAL7 gene via the Gal4-binding sequence, is sufficient for activating GAL7 in the absence of Kin28. We have further found that AR2 has an ability to recruit TATA box-binding protein-associated factors (TAFs) to the promoter. Consistently, transcription from promoters occupied naturally or artificially with TAFs is sustained in the absence of Kin28 function. CONCLUSIONS: These results show that CTM modulates activation function of AR2 in the Hsf1 molecule. We also suggest that recruitment of TAFs to a promoter is involved in the Kin28-independent transcription.
Artificial recruitment of certain Mediator components affects requirement of basal transcription factor IIE.
BACKGROUND: Basal transcription factors are essential for RNA polymerase II (RNAPII)-catalysed transcription of many but not all the mRNA-encoding genes in vivo as well as in vitro. For example, copper-inducible transcription of the copper metallothionein gene CUP1 occurs independently of basal factor TFIIE in budding yeast. To gain insight into the mechanism by which the requirement for TFIIE is bypassed, we artificially recruited certain constituents of Mediator, a large protein complex transmitting signals from various activators to the RNAPII machinery, to the CUP1 promoter by protein fusions with Ace1, the copper-inducible activator. RESULTS: Fusions with Med2 or Pgd1 activated CUP1 independently of TFIIE. Surprisingly, fusions with neither Srb5 nor Med9 circumvented TFIIE requirement for the CUP1 activation. Components of TFIID were similarly recruited to the CUP1 promoter without activation. By using a chromatin immunoprecipitation technique, we found that TFIIE is necessary for stable binding of TFIIH and RNAPII to the ADH1 promoter, whose activation requires TFIIE. However, binding of TFIIH and RNAPII to CUP1 upon its activation did not require TFIIE. CONCLUSIONS: Our results strongly suggest that the TFIIE requirement of a gene is determined by a target(s) in Mediator through which the signal of the cognate activator is transmitted.