Cancer metastasis is a multiple-step process that involves the regulated interaction of diverse cellular proteins. We recently reported that the expression of tumor-associated antigen L6 (TAL6) promoted the invasiveness of lung cancer cells and was inversely correlated with disease-free survival of squamous lung carcinoma patients. We now report that CD13 (aminopeptidase N) can associate with TAL6 and can enhance cancer cell migration. CD13 was shown by coimmunoprecipitation to associate in vitro with TAL6 on several cancer cell lines and to associate in vivo by antibody-mediated copatching immunofluorescence. CD13 was selectively expressed on highly invasive CL1-5 lung cancer cells as compared to poorly invasive CL1-0 lung cancer cells. The role of CD13 aminopeptidase activity in regulating cell motility was investigated with chemical inhibitors, specific antibodies and a catalytically inactive CD13 protein. Inhibition of CD13 aminopeptidase activity by nontoxic concentrations of leuhistin modestly decreased the migration of CL1-5 cells. In contrast, binding of CD13 by specific antibodies significantly reduced both the migration and the invasion of CL1-5 cells. Poorly invasive CL1-0 cells that stably expressed CD13 displayed significantly (p < or = 0.0005) enhanced cell migration (300% of control). Expression of an enzymatically inactive CD13 mutant on CL1-0 cells also significantly (p < or = 0.0005) enhanced cell migration (200% of control). Our results show that TAL6 and CD13 can form a complex on lung cancer cells, that these molecules can modulate cell migration and invasion and that the influence of CD13 on cell motility did not strictly depend on its aminopeptidase activity.

We previously presented YM500, which is an integrated database for miRNA quantification, isomiR identification, arm switching discovery and novel miRNA prediction from 468 human smRNA-seq datasets. Here in this updated YM500v2 database (http://ngs.ym.edu.tw/ym500/), we focus on the cancer miRNome to make the database more disease-orientated. New miRNA-related algorithms developed after YM500 were included in YM500v2, and, more significantly, more than 8000 cancer-related smRNA-seq datasets (including those of primary tumors, paired normal tissues, PBMC, recurrent tumors, and metastatic tumors) were incorporated into YM500v2. Novel miRNAs (miRNAs not included in the miRBase R21) were not only predicted by three independent algorithms but also cleaned by a new in silico filtration strategy and validated by wetlab data such as Cross-Linked ImmunoPrecipitation sequencing (CLIP-seq) to reduce the false-positive rate. A new function ‘Meta-analysis’ is additionally provided for allowing users to identify real-time differentially expressed miRNAs and arm-switching events according to customer-defined sample groups and dozens of clinical criteria tidying up by proficient clinicians. Cancer miRNAs identified hold the potential for both basic research and biotech applications.

The expression pattern of mucin genes was studied in 7 lung adenocarcinoma cell lines (CL1, CL2, CL3, NCL2, PC9, PC13, PC14) and 12 lung adenocarcinoma tissues. CL1 and PC 13 are poorly differentiated cell lines with low mucin glycoprotein production. The other 5 cell lines are well differentiated and produce a higher amount of mucins. Total RNA was extracted from these cell lines. Northern blot analysis was performed by hybridization with specific antisense oligonucleotide probes recognizing mucin-specific tandem repeats of 4 mucin genes (MUC1, MUC2, MUC3, MUC4). RT-PCR was carried out to amplify the 3′ and 5′ nonrepetitive coding regions of MUC1 and the 5’ nonrepetitive coding region of MUC2. All these cell lines expressed MUC1, MUC2, MUC3, and MUC4 mRNA but in variable mounts. The poorly differentiated cell lines (CL1 and PC 13) had a relatively low level of expression of MUC1, MUC2, MUC3 and MUC4. RT-PCR, with primers amplifying the MUC1 nonrepetitive coding region 5′ end, 293 bp, and the 3′ end, 522 bp, as well as the MUC2 nonrepetitive 5′ coding region, 308 bp, revealed the presence of MUC1 and MUC2 mRNA in all the cell lines. Sequence analysis of the PCR products were very homologous, similar to previously published MUC1 and MUC2 cDNA sequences. The expression pattern of mucin genes is consistent with that of mucin glycoproteins as studied using biochemical and immunological methods. Northern blotting and RT-PCR analysis in 12 lung adenocarcinoma tissues with various grades of differentiation (6 poorly differentiated adenocarcinomas and 6 moderately to well-differentiated adenocarcinomas) showed heterogeneous expression of the 4 mucin genes in tissues without clear correlation with the differentiation grade. Therefore the clinical implications of the differential expression of the mucin genes need further investigation.