Novel polysome messages and changes in translational activity appear after induction of adipogenesis in 3T3-L1 cells
1 Institute of Veterinary Medicine, University of Göttingen, Burckhardtweg 2, 37077 Göttingen, Germany
2 Statistical Bioinformatics, Department of Medical Statistics, University Medical Center, Humboldtallee 32, 37073 Göttingen, Germany
3 DNA Microarray Facility Göttingen, Department of Developmental Biochemistry, University of Göttingen, Humboldtallee 23, 37073 Göttingen, Germany
4 Department of Internal Medicine, Metabolic Diseases and Medical Molecular Biology, Paracelsus Private Medical University Salzburg, Müllner Hauptstr. 48, 5020 Salzburg, Austria
BMC Molecular Biology 2012, 13:9 doi:10.1186/1471-2199-13-9Published: 21 March 2012
Control of translation allows for rapid adaptation of the cell to stimuli, rather than the slower transcriptional control. We presume that translational control is an essential process in the control of adipogenesis, especially in the first hours after hormonal stimulation. 3T3-L1 preadipocytes were cultured to confluency and adipogenesis was induced by standard protocols using a hormonal cocktail. Cells were harvested before and 6 hours after hormonal induction. mRNAs attached to ribosomes (polysomal mRNAs) were separated from unbound mRNAs by velocity sedimentation. Pools of polysomal and unbound mRNA fractions were analyzed by microarray analysis. Changes in relative abundance in unbound and polysomal mRNA pools were calculated to detect putative changes in translational activity. Changes of expression levels of selected genes were verified by qPCR and Western blotting.
We identified 43 genes that shifted towards the polysomal fraction (up-regulated) and 2 genes that shifted towards free mRNA fraction (down-regulated). Interestingly, we found Ghrelin to be down-regulated. Up-regulated genes comprise factors that are nucleic acid binding (eIF4B, HSF1, IRF6, MYC, POLR2a, RPL18, RPL27a, RPL6, RPL7a, RPS18, RPSa, TSC22d3), form part of ribosomes (RPL18, RPL27a, RPL6, RPL7a, RPS18, RPSa), act on the regulation of translation (eIF4B) or transcription (HSF1, IRF6, MYC, TSC22d3). Others act as chaperones (BAG3, HSPA8, HSP90ab1) or in other metabolic or signals transducing processes.
We conclude that a moderate reorganisation of the functionality of the ribosomal machinery and translational activity are very important steps for growth and gene expression control in the initial phase of adipogenesis.