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Figure 8


Fig. 8. HIF1{alpha} directly regulates Sox9 expression. (A) Increase in Sox9 and Pgk1 mRNA level upon infection of mesenchymal cells with lentivirus encoding stabilized human HIF1{alpha} (n=2). (B,C) Substantial reduction in Sox9 mRNA and protein level in AdCre-infected cells was detected by quantitative RT-PCR analysis (B) and immunoblot assay (C). (D,E) Quantitative RT-PCR analyses for Sox6 (D) and Sox5 (E) show a marked decrease in AdCre-infected cells under hypoxia (n=3). (F) Schematic view of 3.0 kb Sox9 promoter region showing the position of four potential HIF1{alpha} binding sites relative to the transcription start site. (G,H) Chromatin immunoprecipitation assay of mouse Sox9 gene. Real-time PCR quantification of Hif1{alpha} binding to regions A-D (amplicons) is indicated by the percentage of total input chromatin DNA (n=2). (I) Immunoprecipitation assay with plasmids that contain control Sox9 promoter or plasmids that contain the Sox9 promoter with a four-nucleotide substitution in the HRE 398 (n=2).





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