Molecular cloning, identification and analysis of lung squamous cell carcinoma-related genes

Abstract

Objective: To clone and identify genes differentially expressed in human lung squamous cell carcinoma (LSCC). Methods: A subtracted cDNA library of human LSCC was constructed by using suppression subtractive hybridization (SSH) method. Through screening, the subtracted library clones, representing mRNAs that are truly differentially expressed in LSCC but not in normal lung tissues, were selected out to identify by semi-quantitative RT-PCR in 12 patients of LSCC and performed DNA sequencing. Nucleic acid homology searches were performed using the BLAST program. Partial novel genes were detected by Northern blot. Results: By this technique, we obtained 10 differentially expressed gene cDNA fragments of LSCC. Among them six were already known genes; two sequences were already identified but their functions were still unknown (hypothetical protein); two were novel (GenBank accession number were AF363068 and AY032661, respectively). The results from semi-quantitative RT-PCR showed that the transcription expression level of these clones including PPP1CB, caluminin, S100A2, HSNOV1, OCIA and AY032661 was down-regulated in 12 cases of LSCC, while the transcription of HSP90, ferritin, gp96 and AF363068 was up-regulated in same cases. Conclusion: SSH is a powerful technique of high sensitivity for the detection of differential gene expression in LSCC and an effective method to clone novel genes. Six already known genes identified by SSH technique have been already implicated in the pathogenesis of lung carcinogenesis, or they are involved in immunological defense mechanism in human body. Two hypothetical proteins probably also play an important role in lung cancer pathogenesis. The function of two novel genes in lung carcinogenesis is under research.