Research Paper Volume 6, Issue 7 pp 587—601

Figure 1. Thiamine supply is activated by Phx1. (A) The mRNA levels of genes involved in thiamine biosynthesis (nmt1⁺, nmt2⁺), transport (bsu1⁺, thi9⁺), and metabolism (tnr3⁺) in wild type (WT; JH43) and Δphx1(ESX5) mutant. Cells were grown in minimal media to either exponential or stationary phases, with (A2) or without (A1) adding 10 μM thiamine. The gene-specific mRNA levels were measured by qRT-PCR, along with that of act1⁺ mRNA as an internal control. Each internally normalized expression level at stationary phase was presented in the figure as a relative value to the level in exponential cells. Average values from three independent experiments were presented with standard deviations. (B, C) Intracellular levels of total thiamine pool (B) and TPP (C). Wild-type and Δphx1 mutant cells grown in minimal media were harvested at early exponential (12 h), late exponential (36 h), and late stationary (80 h) phases. Thiamine and thiamine phosphates (TMP, TPP) were extracted and measured by HPLC. Total thiamine is the sum of thiamine and thiamine phosphates. Average values with standard deviations (error bars) from at least three independent experiments were presented. Asterisks (**) represents p-value of <0.05 in Student t-test.