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Fig. 3 | Genome Biology

Fig. 3

From: A post-transcriptional program of chemoresistance by AU-rich elements and TTP in quiescent leukemic cells

Fig. 3

Phosphorylation of TTP stabilizes ARE-bearing TNFα in G0 leukemic cells. a Boxplot of ARE scores (SI methods) in the 3′UTRs of genes which are up- or downregulated at the translatome or RNA levels in G0 compared to S+ cells. b Venn diagram of genes that are upregulated at the translatome level and contain AREs (left) and examples of such genes (right). See also Additional file 3: Table S2 for a full list of genes. c Expression of ARE genes at the RNA and translatome levels. d Scatter plot showing the expression of RNA-binding protein genes from RBPDB database (SI methods). TTP is indicated with a green dot. e Western analysis of TTP in lysates from multiple leukemic cell lines in the absence or presence of alkaline phosphatase (AP). Phospho-TTP is indicated with an arrow. f Bar graph shows TNFα mRNA expression normalized to GAPDH mRNA upon overexpression of vector or c-myc tagged non-phosphorylatable mutant TTP (TTP-AA) in AraC-treated THP1 or K562 cells. Western analysis of TTP-AA with c-myc antibody (right). g Half-life of TNFα mRNA. TTP-deficient BMDM cells were transduced with doxycycline inducible plasmids that express GFP vector, TTP wild-type, or TTP-AA mutant. Cells were induced with 1 μg/ml doxycycline prior to 1 μM AraC treatment. Western analysis of induction of TTP protein. TNFα mRNA level was measured at indicated time points by qPCR after transcriptional arrest with 5 μg/ml actinomycin D treatment. h Association of TTP-AA with TNFα mRNA in AraCS cells. TTP-AA was immunoprecipitated with GFP antibody from AraC-treated BMDM cells expressing GFP-tagged TTP-AA (Western blot), followed by qPCR analysis of TNFα mRNA (graph). *p ≤ 0.05. Data are represented as average ± SEM. See also Additional file 1: Figure S3 and Additional file 3: Table S2

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