Genome-wide identification and expression analysis of microRNA involved in small cell lung cancer via deep sequencing

Abstract

Abstract. Small cell lung cancer is a major cause of mortality worldwide. microRNAs (miRNAs) are involved in various biological processes through regulating gene expression. In the present study, to identify the miRNAs involved in human small cell lung cancer at the genome-wide level, Solexa sequencing was employed to sequence two small RNA (sRNA) libraries from small cell lung cancer tissues (LC sRNA library) and the corresponding normal tissues (NT sRNA library). Deep sequencing of the two sRNA libraries identified a number of conserved miRNAs and differential expression analysis of these miRNAs revealed 81 miRNAs differentially expressed in small cell lung cancer, of which more than half were downregulated. The expression trends determined by sequencing were validated by reverse transcription-quantitative polymerase chain reaction analysis. The annotations for the targets of these miRNAs were predicted. This study provides valuable information for understanding the regulatory mechanisms of miRNAs involved in human small cell lung cancer. IntroductionMicroRNAs (miRNAs) are small non-coding RNA molecules, ~22 nucleotides long, which are widely distributed in plants and animals (1,2). The primary miRNA transcripts are cleaved by Drosha and Dicer enzymes to form mature miRNAs, which serve as posttranscriptional negative gene regulators by either cleaving mRNA or inhibiting translation (3,4). Through such mechanisms, miRNAs are involved in various biological processes, including organ development, tissue differentiation, cell cycle regulation and cancer development (4-6).Lung cancer is the predominant cause of cancer-related mortality worldwide and its pathogenesis is closely associated with tobacco smoking. Lung cancer is categorized into two main histological groups: Non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC). Regulation of miRNAs is closely associated with tumor initiation, promotion and progression (7). Recently, increasing evidence has demonstrated that miRNAs may be associated with lung cancer. A previous study revealed that an increase in hsa-miR-196a expression levels was a characteristic molecular change in NSCLC (8,9), and that hsa-miRNA-196a promoted NSCLC cell proliferation and invasion via targeting HOXA5 (10). Overexpression of miR-200 was shown to reduce the expression levels of NSCLC prognostic biomarkers in H1299 and BEAS-2B cells (11). miR-449a, which inhibited migration and invasion through targeting c-Met, was found to be downregulated in NSCLC tissues and cell lines (12). Reduced expression of miR-101 was associated with overexpression of EZH2 and exhibited tumor-suppressive functions in NSCLC (13). This evidence suggests that miRNAs are crucial in NSCLC; however, little is known regarding the functions and regulation of miRNAs in SCLC.In the present study, Solexa sequencing was used to generate a large quantity of small RNA (sRNA) data for SCLC and corresponding normal tissues. Sequence analysis was employed to identify specific miRNAs associated with SCLC and these ...