Function-Based Discovery of Significant Transcriptional Temporal Patterns in Insulin Stimulated Muscle Cells

Abstract

BackgroundInsulin action on protein synthesis (translation of transcripts) and post-translational modifications, especially of those involving the reversible modifications such as phosphorylation of various signaling proteins, are extensively studied but insulin effect on transcription of genes, especially of transcriptional temporal patterns remains to be fully defined.Methodology/Principal FindingsTo identify significant transcriptional temporal patterns we utilized primary differentiated rat skeletal muscle myotubes which were treated with insulin and samples were collected every 20 min for 8 hours. Pooled samples at every hour were analyzed by gene array approach to measure transcript levels. The patterns of transcript levels were analyzed based on a novel method that integrates selection, clustering, and functional annotation to find the main temporal patterns associated to functional groups of differentially expressed genes. 326 genes were found to be differentially expressed in response to in vitro insulin administration in skeletal muscle myotubes. Approximately 20% of the genes that were differentially expressed were identified as belonging to the insulin signaling pathway. Characteristic transcriptional temporal patterns include: (a) a slow and gradual decrease in gene expression, (b) a gradual increase in gene expression reaching a peak at about 5 hours and then reaching a plateau or an initial decrease and other different variable pattern of increase in gene expression over time.Conclusion/SignificanceThe new method allows identifying characteristic dynamic responses to insulin stimulus, common to a number of genes and associated to the same functional group. The results demonstrate that insulin treatment elicited different clusters of gene transcript profile supporting a temporal regulation of gene expression by insulin in skeletal muscle cells.

p90 Ribosomal S6 kinases play a significant role in early gene regulation in the cardiomyocyte response to Gq-protein-coupled receptor stimuli, endothelin-1 and α1-adrenergic receptor agonists

“…In cardiomyocytes exposed to ET-1 [14] or α 1 -AR agonists (Figures 4 and 6), regulation of early gene expression occurs according to temporal profiles with many RNAs exhibiting only transient expression. This occurs in other systems including (as recent examples) skeletal muscle exposed to insulin [39] and macrophages exposed to pro-inflammatory stimuli [40]. Given the difference in time point studied in our experiments (1 h) and in the cancer cell system (24 h), and the nature of the two cell types, it is perhaps not surprising that there is little overlap in the RSK-responsive genes.…”

Reverse-engineering the genetic circuitry of a cancer cell with predicted intervention in chronic lymphocytic leukemia

“…So, at each time point after stimulation, we studied gene expression both in stimulated cells and in control (unstimulated) cells. Given that several temporal gene expression profiles have revealed complex gene expression after cellular stimulation (3,20), we considered that genes with both high expression level and those with a specific expression pattern (regardless of their expression level) are relevant in the program (21). Gene selection methods based upon selection of highly differentially expressed genes are widely used.…”

Insulin-dependent transcriptional control in L6 rat myotubes is associated with modulation of histone acetylation and accumulation of the histone variant H2A.Z in the proximity of the transcriptional start site

“…This study has a limitation, as we focused our investigation on a selection of insulin-regulated genes (Culbert and Tavare 2002;Zhang et al 2001, Di Camillo et al 2012. Further studies employing genome-wide approaches are warranted.…”