Integrated mRNA and microRNA transcriptome sequencing characterizes sequence variants and mRNA-microRNA regulatory network in nasopharyngeal carcinoma model systems

FEBS Open Bio. 2014 Jan 13;4:128-40. doi: 10.1016/j.fob.2014.01.004. eCollection 2014.

Abstract

Nasopharyngeal carcinoma (NPC) is a prevalent malignancy in Southeast Asia among the Chinese population. Aberrant regulation of transcripts has been implicated in many types of cancers including NPC. Herein, we characterized mRNA and miRNA transcriptomes by RNA sequencing (RNASeq) of NPC model systems. Matched total mRNA and small RNA of undifferentiated Epstein-Barr virus (EBV)-positive NPC xenograft X666 and its derived cell line C666, well-differentiated NPC cell line HK1, and the immortalized nasopharyngeal epithelial cell line NP460 were sequenced by Solexa technology. We found 2812 genes and 149 miRNAs (human and EBV) to be differentially expressed in NP460, HK1, C666 and X666 with RNASeq; 533 miRNA-mRNA target pairs were inversely regulated in the three NPC cell lines compared to NP460. Integrated mRNA/miRNA expression profiling and pathway analysis show extracellular matrix organization, Beta-1 integrin cell surface interactions, and the PI3K/AKT, EGFR, ErbB, and Wnt pathways were potentially deregulated in NPC. Real-time quantitative PCR was performed on selected mRNA/miRNAs in order to validate their expression. Transcript sequence variants such as short insertions and deletions (INDEL), single nucleotide variant (SNV), and isomiRs were characterized in the NPC model systems. A novel TP53 transcript variant was identified in NP460, HK1, and C666. Detection of three previously reported novel EBV-encoded BART miRNAs and their isomiRs were also observed. Meta-analysis of a model system to a clinical system aids the choice of different cell lines in NPC studies. This comprehensive characterization of mRNA and miRNA transcriptomes in NPC cell lines and the xenograft provides insights on miRNA regulation of mRNA and valuable resources on transcript variation and regulation in NPC, which are potentially useful for mechanistic and preclinical studies.

Keywords: AIP, aryl hydrocarbon receptor interacting protein; BAX, BCL2-asscoiated X protein; CIITA, class II, major histocompatibility complex, transactivator; DKK1, Dickkopf-Like protein 1; EBV, Epstein–Barr virus; ECM, extracellular matrix; EGFR, epidermal growth factor receptor; EGR1, early growth response 1; FBLN2, fibulin 2; GADD45, growth arrest and DNA-damage-inducible; GNG11, guanine nucleotide binding protein (G protein), Gamma 11; GO, gene ontology; GSTP1, glutathione S-transferase pi 1; IL18, interleukin 18; INDEL, insertion and deletion; LMP1, Epstein–Barr virus latent membrane protein 1; LTBP2, latent transforming growth factor beta binding protein 2; MDM2, MDM2 oncogene, E3 ubiquitin protein ligase; MET, met proto-oncogene; MMP19, matrix metallopeptidase 19; NGS, next-generation sequencing; NPC, nasopharyngeal carcinoma; Nasopharyngeal carcinoma; Nasopharyngeal cell lines/xenograft (NP460, HK1, C666, X666); PI3K, phosphoinositide 3-kinase; PTEN, phosphatase and tensin homolog; RNA sequencing; RNASeq, RNA sequencing; SNP, single nucleotide polymorphism; TNFRSF9, tumour necrosis factor receptor superfamily, member 9; TP53; Transcriptome analysis; UTR, untranslated region; miRNA, microRNA.