Microarray analysis identifies a common set of cellular genes modulated by different HCV replicon clones - PubMed (original) (raw)

Microarray analysis identifies a common set of cellular genes modulated by different HCV replicon clones

Anna Rita Ciccaglione et al. BMC Genomics. 2008.

Abstract

Background: Hepatitis C virus (HCV) RNA synthesis and protein expression affect cell homeostasis by modulation of gene expression. The impact of HCV replication on global cell transcription has not been fully evaluated. Thus, we analysed the expression profiles of different clones of human hepatoma-derived Huh-7 cells carrying a self-replicating HCV RNA which express all viral proteins (HCV replicon system).

Results: First, we compared the expression profile of HCV replicon clone 21-5 with both the Huh-7 parental cells and the 21-5 cured (21-5c) cells. In these latter, the HCV RNA has been eliminated by IFN-alpha treatment. To confirm data, we also analyzed microarray results from both the 21-5 and two other HCV replicon clones, 22-6 and 21-7, compared to the Huh-7 cells. The study was carried out by using the Applied Biosystems (AB) Human Genome Survey Microarray v1.0 which provides 31,700 probes that correspond to 27,868 human genes. Microarray analysis revealed a specific transcriptional program induced by HCV in replicon cells respect to both IFN-alpha-cured and Huh-7 cells. From the original datasets of differentially expressed genes, we selected by Venn diagrams a final list of 38 genes modulated by HCV in all clones. Most of the 38 genes have never been described before and showed high fold-change associated with significant p-value, strongly supporting data reliability. Classification of the 38 genes by Panther System identified functional categories that were significantly enriched in this gene set, such as histones and ribosomal proteins as well as extracellular matrix and intracellular protein traffic. The dataset also included new genes involved in lipid metabolism, extracellular matrix and cytoskeletal network, which may be critical for HCV replication and pathogenesis.

Conclusion: Our data provide a comprehensive analysis of alterations in gene expression induced by HCV replication and reveal modulation of new genes potentially useful for selection of antiviral targets.

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Figures

Figure 1

Figure 1

(A) Venn diagram of probes differentially expressed in 21-5 vs. 21-5c (dataset 1) and 21-5 vs. Huh-7 (dataset 2) comparisons. The number of probes differentially expressed in both comparisons was 156, and 104 (7.7%) out of 1344 total probes showed concordant expression (up-regulated or down-regulated in both datasets). (B) Venn diagram of the differentially expressed probes in 21-5 vs. 21-5c (dataset 1) and HCV clones vs. Huh-7 (dataset 3) comparisons. The number of probes differentially expressed in both comparisons was 88, and 58 (4.4%) out of 1327 total probes showed concordant expression. (C) Venn diagram of the 104 and 58 probes identified 39 (31,7%) common probes out of 123.

Figure 2

Figure 2

Scatterplot comparison of the log2 ratio of the 58 probes altered in the indicated datasets. Log2 ratios of genes from dataset 3 are plotted on the y-axis and from dataset 1 on the x-axis.

Figure 3

Figure 3

Real-time PCR validation of the microarray data, performed for 7 genes modulated by HCV. Total RNA from the 21-5 and 21-5 cured cell lines was used to assess mRNA levels using real time RT-PCR. Levels were normalized to cellular GAPDH; mRNA levels from 21-5 cured cells were set as the basis for the comparative results. Shown are the means (± SD) of three independent experiments. Fold-changes calculated for the microarray data are also indicated.

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