Abstract
Yeast genes involved in thiamin pyrophosphate synthesis (THI genes) are transcriptionally induced in response to thiamin starvation. In this system, three proteins (Thi2, Thi3p, Pdc2) act as positive regulatory factors. Thi2 is a DNA-binding protein whose target consensus DNA sequence in the upstream region of THI is deduced. Thi3 acts as a thiamin pyrophosphate sensor and upregulates THI expression when thiamin pyrophosphate is not bound to Thi3. We found that Thi3 is associated with Pdc2 directly and to a lesser extent with Thi2 and that these interactions are partially disturbed by thiamin pyrophosphate. We also demonstrated that Pdc2 transactivates gene expression and interacts with the upstream region of THI genes, both of which are enhanced by thiamin starvation. These enhancements were not observed in thi2 or thi3 strain. When the C-terminal Thi3-interacting domain of Pdc2 was truncated, Pdc2 expressed strikingly transactivation activity in a Thi3-independent fashion. These results suggested that the recruitment of Pdc2 to the upstream region of THI genes is facilitated by Thi2 bound on its target DNA sequence via interaction with Thi3 and that the ternary Thi2/Thi3/Pdc2 complex transactivates THI genes. It is also proposed that Thi3 causes a conformational change in Pdc2 leading to full transactivation activity.
