Unknown,Transcriptomics,Genomics,Proteomics

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Gut microbiome metabolites alter genomic networks in PC12 cells relevant to autism spectrum diorders

ABSTRACT: Alterations in the composition of the gut microbiome have an emerging role in brain function and behaviour. We have porposed that short chain fatty acids (SCFA) including propionate and butyrate which are present in the diet and are fermantation products of many gastrointestinal bacteria are contributing environmental factors in autism spectrum disorders (ASD). Here we used the microarray technology to compare global changes in gene expression profiles following exposure of PC12 cells to structurally related SCFA propionate and butyrate each in two different concentrations. Large number of affected genes, common for both SCFA were identified, including genetic networks and GO processes implicated in ASD. PC12 cells were exposed to propionate or butyrate. RNA was isolated from each experimental group (n=6, pooled samples) and subjected to genome-wide expression profiling using Affymetrix microarrays to reveal dose- and SCFA-specific changes in gene expression and specific molecular pathways or processes affected as compared to vehicle treated controls.

ORGANISM(S): Rattus norvegicus

SUBMITTER: Bistra Nankova

PROVIDER: E-GEOD-56516 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Background: Acetate, propionate, and butyrate are the main short-chain fatty acids (SCFA) produced in the colon as a result of microbial fermentation of dietary fibers. An increasing amount of evidence suggests that these SCFA have major health benefits. The composition of the microbiota is altered by dietary fat, and this is believed to impact SCFA production. Currently it is unknown whether host gene expression responses to SCFA are modulated by fat content of the diet. The aim of this study was to compare the changes in colonic gene expression profiles after acetate, propionate and butyrate infusions between a low fat and high fat diet. Methods: Male C57BL/6J mice were fed semi-synthetic low fat (10 energy%) or high fat (45 E%) diets starting 2 weeks before the SCFA treatment period. During treatment, mice received a rectal infusion of either an acetate, propionate, butyrate, or a saline (control) solution for 6 consecutive days, after which colon was subjected to gene expression profiling. Unsupervised visualization of the dataset was performed using Independent Principal Component Analysis. For each SCFA, similarities of its effects on a low fat and a high fat diet were assessed using Rank-Rank Hypergeometric Overlap. In addition, differentially expressed genes were identified, and gene set enrichment analysis was performed to determine functional implications of the regulated genes. Results: Taking into account the complete dataset, we observed that more variation in gene expression profiles was explained by fat content of the diet than by SCFA treatment. Gene expression responses to acetate and butyrate were similar on the low fat versus high fat diet, but were opposite for propionate. Functionally the expression changes reflected differential modulation of several metabolic processes; genes involved in oxidative phosphorylation, lipid catabolism, lipoprotein metabolism and cholesterol transport were suppressed by acetate and butyrate treatment, whereas propionate treatment resulted in changes in fatty acid and sterol biosynthesis, and in amino acid and carbohydrate metabolism. Conclusions: We demonstrated that dietary fat content impacts the colonic gene expression response to propionate, and to a lesser extent to acetate and butyrate. The study demonstrates that knowledge on diet composition is essential when studying effects of SCFAs on metabolism.

Project description:The nerve growth factor NGF has been shown to cause cell fate decisions towards either differentiation or proliferation depending on the relative activity of downstream pERK, pAKT or pJNK signaling. However, how these protein signals are translated into and fed back from transcriptional activity to complete cellular differentiation over a time span of hours to days is still an open question. Using dynamic proteome and transcriptome data from NGF-stimulated PC12 cells over a time span of 24 hours we inferred a dynamic Boolean model capturing the temporal sequence of protein signaling, transcriptional response and subsequent autocrine feedback. Optimal model topology was achieved by introducing multiple time scales for fast cellular signaling and slower gene response and subsequent fitting to the experimental data. The integrated model confirmed the parallel use of MEK/ERK, AKT/PI3K and JNK/JUN for PC12 cell differentiation. Redundancy of cell signaling is demonstrated from the inhibition of the different MAPK pathways. As suggested in silico and confirmed in vitro, differentiation was substantially suppressed under JNK/JUN inhibition, yet delayed only under MEK/ERK inhibition. Most importantly, we found that sustained transcriptional feedback is necessary to induce bistability in the cell fate switch. De novo Gene expression was necessary to activate autocrine feedback that caused integrin signaling to perpetuate the MAPK activity, finally resulting in the expression of late, differentiation related genes. Thus, the cellular decision towards differentiation depends on the establishment of a transcriptome-induced positive feedback between protein signaling and gene expression thereby constituting a robust control between proliferation and differentiation. To eludicate the transcriptome response of PC12 cells time-resolved gene expression data of NGF or EGF simulated PC12 were recorded at t = [0.5h 1h, 2h, 3h, 4h, 5h, 6h, 8h, 12h, 24h, 48h] for NGF and at t=[1h, 2h, 3h, 4h, 6h, 8h, 12h, 24h]. Control time points of unstimulated PC12 cells were taken at t=[0h, 2h, 4h, 6h, 8h, 24h, 48h]. Additionally, the MEK-inhibitor UO126 was added together with NGF and the gene response was measured at t=[1h, 2h, 4h, 6h, 8h, 12h, 24h, 48h] and for UO126 alone at t=[1h, 2h, 4h, 8h, 12h, 48h]. Total RNA was isolated, labeled and hybridized to an Illumina ratRef-12 bead array (Illumina, San Diego, CA, USA) according to the manufacturerâ??s protocol.

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Gut microbiome metabolites alter genomic networks in PC12 cells relevant to autism spectrum diorders

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