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FIGURE 5. Bteb1 and TrβA mRNAs are up-regulated by T3, and BTEB1 associates with the proximal TrβA promoter in XTC-2 cells. A, T3 up-regulates Bteb1 mRNA in XTC-2 cells with faster kinetics than TRβ. XTC-2 cells were treated with T3 (5 nM) for various times before harvest for RNA isolation and semi-quantitative RT-PCR analysis of Bteb1 and TrβA mRNA expression. Gene expression was normalized to the level of rpL8 expression (a housekeeping gene). Bteb1 mRNA was maximally induced at 3 h (p = 0.009; Scheffe's test) and maintained through 48 h of treatment. TrβA mRNA was significantly induced at 6 h (p = 0.001), reached a maximum by 12 h, and was maintained through 48 h. Bars represent the mean ± S.E. (n = 6 wells/time point), and letters above the means indicate significant differences among time points (i.e. means with the same letter are not significantly different; p < 0.05; Scheffe's test). B, BTEB1 associates with the proximal TrβA promoter in XTC-2 cells. XTC-2 cells were treated with T3 (5 nM) for 24 h, and we used ChIP assay coupled with quantitative real time PCR to detect BTEB1 association with the TrβA gene. We found significantly greater association of BTEB1 at an upstream region of the promoter (overlapping with regions A and B shown in Fig. 2; -885 to -752), which contains multiple GC boxes compared with a region in the 5′-UTR (region G; +166 to +322) that has only one GC box, or the exon 5 of the TrβA gene which has no GC boxes. Letters indicate significant differences among gene regions (i.e. means with the same letter are not significantly different; p < 0.05; Scheffe's test). C, treatment of XTC-2 cells with T3 (5 nM, 24 h) increases BTEB1 association with the upstream TRβA promoter (region A/B) as analyzed by ChIP assay (*, p = 0.043; t test). Image published in: Bagamasbad P et al. (2008) Copyright © 2008. Image reproduced with permission of the publisher and the copyright holder. This is an Open Access article distributed under the terms of the Creative Commons Attribution License. Larger Image Printer Friendly View

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