No abstract

BackgroundA pool of small RNA fragments (RFs) derived from diverse cellular RNAs has recently emerged as a rich source of functionally relevant molecules. Although their formation and accumulation has been connected to various stress conditions, the knowledge on RFs produced upon viral infections is very limited. Here, we applied the next generation sequencing (NGS) to characterize RFs generated in the hepatitis C virus (HCV) cell culture model (HCV-permissive Huh-7.5 cell line).ResultsWe found that both infected and non-infected cells contained a wide spectrum of RFs derived from virtually all RNA classes. A significant fraction of identified RFs accumulated to similar levels as miRNAs. Our analysis, focused on RFs originating from constitutively expressed non-coding RNAs, revealed three major patterns of parental RNA cleavage. We found that HCV infection induced significant changes in the accumulation of low copy number RFs, while subtly altered the levels of high copy number ones. Finally, the candidate RFs potentially relevant for host-virus interactions were identified.ConclusionsOur results indicate that RFs should be considered an important component of the Huh-7.5 transcriptome and suggest that the main factors influencing the RF biogenesis are the RNA structure and RNA protection by interacting proteins. The data presented here significantly complement the existing transcriptomic, miRnomic, proteomic and metabolomic characteristics of the HCV cell culture model.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-017-3891-3) contains supplementary material, which is available to authorized users.

Activation-induced cytidine deaminase (AID) is known for its established role in antibody production. AID induces the diversification of antibodies by deaminating deoxycytidine (C) within immunoglobulin genes. The capacity of AID to deaminate 5-methyldeoxycytidine (5 mC) and/or 5-hydroxymethyldeoxycytidine (5 hmC), and consequently AID involvement in active DNA demethylation, is not fully resolved. For instance, structural determinants of AID activity on different substrates remain to be identified. To better understand the latter issue, we tested how mutations in human AID (hAID) influence its ability to deaminate C, 5 mC, and 5 hmC in vitro. We showed that each of the selected mutations differentially affects hAID’s ability to deaminate C and 5 mC. At the same time, we did not observe hAID activity on 5 hmC. Surprisingly, we found that the N51A hAID mutant, with no detectable activity on C, efficiently deaminated 5 mC, which may suggest different requirements for C and 5 mC deamination. Homology modeling and molecular dynamics simulations revealed that the pattern of enzyme-substrate recognition is one of the important factors determining enzyme activity on C and 5 mC. Consequently, we have proposed mechanisms that explain why wild type hAID more efficiently deaminates C than 5 mC in vitro and why 5 hmC is not deaminated.

Hepatitis C virus (HCV) infection is one of the major causes of chronic liver diseases. Unfortunately, the mechanisms of HCV infection-induced liver injury and host-virus interactions are still not well recognized. To better understand these processes we determined the changes in the host gene expression that occur during HCV infection of Huh-7.5 cells. As a result, we identified genes that may contribute to the immune and metabolic cellular responses to infection. Pathway enrichment analysis indicated that HCV induced an increased expression of genes involved in mitogen-activated protein kinases signaling, adipocytokine signaling, cell cycle and nitrogen metabolism. In addition, the enrichment analyses of processes and molecular functions revealed that the up-regulated genes were mainly implicated in the negative regulation of phosphorylation. Construction of the pathway-gene-process network enabled exploration of a much more complex landscape of molecular interactions. Consequently, several essential processes altered by HCV infection were identified: negative regulation of cell cycle, response to endoplasmic reticulum stress, response to reactive oxygen species, toll-like receptor signaling and pattern recognition receptor signaling. The analyses of genes whose expression was decreased upon HCV infection showed that the latter were engaged in the metabolism of lipids and amino acids. Moreover, we observed disturbance in the cellular antiviral defense. Altogether, our results demonstrated that HCV infection elicits host response that includes a very wide range of cellular mechanisms. Our findings significantly broaden the understanding of complex processes that accompany HCV infection. Consequently, they may be used for developing new host-oriented therapeutic strategies.

Copy number variation (CNV) is a newly discovered form of intra-species genetic polymorphism that is defined as deletions or duplications of genome segments ranging from 1 kbp to several Mbp. CNV accounts for the majority of the genetic variation observed in humans (CNV regions cover more than 10% of the human genome); therefore, it may significantly influence both the phenotype and susceptibility to various diseases. Unfortunately, the impact of CNV on a number of diseases, including hepatitis C virus (HCV) infection, remains largely unexplored. Here, we analyzed 421 human genes encoding proteins that have been shown to interact with HCV proteins or genomic RNA (proteins from the HCV-human interactome). We found that 19 of the 421 candidate genes are located in putative CNV regions. For all of these genes, copy numbers were determined for European, Asiatic and African populations using the multiplex ligation-dependent amplification (MLPA) method. As a result, we identified 4 genes, IGLL1, MLLT4, PDPK1, PPP1R13L, for which the CN-genotype ranged from 1 to 6. All of these genes are involved in host-virus interaction; thus, their polymorphism has a potential impact on the development of HCV infection and/or therapy outcome.