Dynamics of mRNA and polysomal abundance in early 3T3-L1 adipogenesis

Abstract

Background: Adipogenesis is a complex process, in which immature pre-adipocytes change morphology, micro-anatomy and physiology to become mature adipocytes. These store and accumulate fat and release diverse hormones. Massive changes in protein content and protein composition of the transforming cell take place within a short time-frame.In a previous study we analyzed changes in the abundance of free and polysomal, i.e. ribosome bound, RNAs in the first hours of adipogenesis in the murine cell line 3T3-L1. Here we analyze changes of mRNA levels and their potential contribution to the changing protein pool by determination of mRNA levels and ribosome binding to mRNAs in 3T3-L1 cells stimulated for adipogenesis. We grouped mRNA species into categories with respect to up- or down-regulated transcription and translation and analyzed the groups regarding specific functionalities based on Gene Ontology (GO).

Results: A shift towards up-regulation of gene expression in early adipogenesis was detected. Genes up-regulated at the transcriptional (TC:up) and translational (TL:up) level (TC:up/TL:up) are very likely involved in control and logistics of translation. Many of them are known to contain a TOP motif. In the TC:up/TL:unchanged group we detected most of the metal binding proteins and metal transporters. In the TC:unchanged/TL:up group several factors of the olfactory receptor family were identified, while in TC:unchanged/TL:down methylation and repair genes are represented. In the TC:down/TL:up group we detected many signaling factors. The TC:down/TL:unchanged group mainly consists of regulatory factors.

Conclusions: Within the first hours of adipogenesis, changes in transcriptional and translational regulation take place. Notably, genes with a specific biological or molecular function tend to cluster in groups according to their transcriptional and translational regulation.

Figure 1

Changes in transcriptional and translational activity. The smooth scatter plot shows the genes of groups 1–9 in Table 1 with some selected outliers being highlighted.

Figure 2

GO term visualization (BP) for gene groups of interest. The scatterplot shows the terms after redundancy reduction via REViGO, representing functional clusters. The bubble color indicates the p-values of the topGO analysis, and the bubble size indicates the frequency of the GO term in the underlying GOA database, i.e. the more general the term, the larger the bubbles. The GO terms cluster together in the semantic space according to functional similarity, without intrinsic meaning of semantic space units. A: TC:up/TL:up (“translation group”), B: TC:up/TL:down (“thrombospondin group”), C: TC:unchanged/TL:down (“methylation and repair group”), D: TC:down/TL:unchanged (“regulation group”).

Figure 3

GO term visualization (MF) for gene groups of interest. The scatterplot shows the terms after redundancy reduction via REViGO, representing functional clusters. The bubble color indicates the p-values of the topGO analysis, and the bubble size indicates the frequency of the GO term in the underlying GOA database, i.e. the more general the term, the larger the bubbles. The GO terms cluster together in the semantic space according to functional similarity, without intrinsic meaning of semantic space units. A: TC:up/TL:unchanged (“metal group”), B: TC:unchanged/TL:up (“olfactory group”), C: TC:down/TL:up (“signaling group”), D: TC:down/TL:down (“Egfr and Ghrl group”).

Figure 4

Scheme of affected cellular processes. The scheme of the adipocyte visualizes prominent cellular processes that are over-represented in the eight analyzed groups. The structures and name of the corresponding group have same colors. The TC:up/TL:up (“translation”) group is represented by ribosomes. The TC:up/TL:unchanged (“metal”) group is symbolized by zinc (channels). The TC:up/TL:down (“thrombospondin”) group is represented by thrombospondin binding to a receptor. Analogously, we chose receptor symbols for the TC:unchanged/TL:up (“olfactory”) group and the TC:down/TL:down (“Egfr and Ghrl”) group. The yellow arrow symbolizes signaling of the TC:down/TL:up (“signaling”) group, e.g. via phosphorylation cascades as initialized by receptor dimerization. The TC:unchanged/TL:down (“methylation and repair”) group is denoted by methyl groups attached to the DNA in the nucleus. The TC:down/TL:unchanged (“regulation”) group is represented by a traffic light.