Transforming growth factor beta 1-stimulated clone (TSC) 22 D4 is a novel key transcriptional co-regulator of hepatic metabolism
Background and aims: The transcriptional co-regulators TBL1/TBLR1 are key metabolic players, controlling liver and serum triglyzeride metabolism. Our aim was to screen for potential novel interaction partners and define their role in metabolism.
Methods: Using luminescence-based mammalian Interactome mapping, we screeneed for TBL1/TBLR1 interaction partners. Adenoviruses expressing a TSC22D4 or a non-specific shRNA were generated for genetic knockdown of TSC22D4. For genetic overexpression of TSC22D4 we established adenoviruses expressing TSC22D4 cDNA under the control of the CMV promoter. For long-term knockdown studies of TSC22D4 we further generated AAVs encoding control or TSC22D4-specific miRNAs under the control of a hepatocyte-specific promoter. To identify novel pathways involved in TSC22D4 metabolism we performed Chip-Sequencing using a flag-tagged TSC22D4 overexpression construct. Gene expression profiling was performed on liver extracts from control or TSC22D4 knockdown mice and primary hepatocytes treated with control or TSC22D4 shRNA adenovirus.
Results: Using luminescence-based mammalian interactome mapping, we identified the transcriptional co-factor TSC22D4 as a novel interaction partner of the transcriptional co-regulators TBL1 and TBLR1. This interaction could be reproduced in a GST pulldown assay. Microarray analysis of shRNA-mediated gene knockdown of TSC22D4 in primary mouse hepatocytes showed a significant induction of key metabolic genes, amongst them fatty acid biosynthesis. Low-fat vs. high-fat feeding over 12 weeks resulted in a significant reduction of TSC22D4 levels in the high-fat diet group, implying a role of TSC22D4 in dietary metabolism. shRNA-mediated as well as miRNA-mediated knockdown of TSC22D4 in C57Bl6 mice resulted in the expression of key metabolic genes; e.g. in the lipogenic pathway and significantly altered liver TG levels as well as serum TG levels. Many of these effects could be reversed using cDNA overexpression of TSC22D4.
Discussion: We identified the transcriptional co-regulator TSC22D4 to interact with the key metabolic regulator TBL1/TBLR1. Genetic knockdown as well as overexpression of TSC22D4 resulted in significant metabolic effects.
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