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Figure 4 | Genome Biology

Figure 4

From: A TATA binding protein regulatory network that governs transcription complex assembly

Figure 4

Testing the TBP regulatory network through genome-wide occupancy and expression profiling. (a) Correlations in RNA and factor occupancy levels, comparing simulated and experimental data. The flux constants governing the core reaction mechanism in Figure 2a were initialized to values specific to clusters 3, 4, 5, 6, 8, and 9 (Additional data file 3). The simulation was run to steady-state. RNA output and the levels of promoter-bound SAGA (Spt3), NC2 (Bur6), Mot1, TFIID (TAF1, TAF5, TAF6, TAF9), and PIC (Rpb1, Rpb2, Rpb7) were reported. These values were then compared to experimentally measured levels of RNA (transcription frequency as defined by [3], and the indicated factor occupancy measured by ChIP-chip [50,52]). Experimental values are median values for each of the six clusters (hence six data points). The x and y axes are on a linear scale. The units for each plot are relative, and thus have no intrinsic meaning except to provide a linear scale demarcating the relative concentration of RNA or promoter-bound species. Correlation coefficients (R) are indicated in the lower right corner of each plot. (b) Correlations between simulated and actual changes in RNA output when the TBP regulatory network is perturbed. The scatter plot reflects changes in actual (from Figure 3) and simulated RNA output relative to a wild-type cell. Median values for each mutant in each of the six clusters are plotted. Cluster 5 data are shown in red. The x and y axes are plotted on a linear scale, with values log2-transformed. The correlation coefficient (R) of all data is shown. (c) Simulated versus actual changes in occupancy of a TBP mutant that is defective in NC2 interactions. Production of HA-tagged TBP(F182V) in a wild-type TBP, taf1(ΔTAND) background was induced with galactose. After 45 minutes, cells were subjected to formaldehyde crosslinking. TBP was immunopurified and Cy3- or Cy5-labeled ChIP DNA was hybridized to yeast intergenic microarrays. The hybridization control reference used galactose-induced HA-tagged wild-type TBP in a taf1(ΔTAND) background, processed in parallel with TBP(F182V). Since the experimental set-up included both endogenous wild-type TBP and TBP(F182V) in the same cell, the simulation simultaneously ran the mechanism in duplicate in which the first mechanism was initialized with parameters for wild-type TBP and the second mechanism with parameters used to simulate the F182V mutation (Additional data file 3). The output being compared is the occupancy level of TBP(F182V) relative to TBP(WT). The line fit and correlation coefficient excludes the outlying cluster 5 data point.

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