Human Microarray Platform Research Articles

IL-10Rα expression is post-transcriptionally regulated by miR-15a, miR-185, and miR-211 in melanoma.

BackgroundIL-10 is an immunoregulatory cytokine that increases during malignant diseases. The purpose of this study was to: i) determine the mRNA amounts of IL-10, IL-10Rα, and IL-10Rβ in cutaneous and uveal melanoma cells and specimens; ii) evaluate their post-transcriptional regulation by miRNAs; iii) ascertain whether miRNA dysregulation may affect IL-10-induced proliferation.MethodsGenome-wide miRNA expression profiling was performed using a human microarray platform. The reference gene mRNA was measured through qPCR. miRNAs/mRNAs interactions were predicted by TargetScan, microRNA, and PITA. Transfections of specific miRNA mimics/inhibitors were carried out. Cell proliferation was assessed by MTT assay in the presence of IL-10 after transfection with miRNA mimics/inhibitors.ResultsThere were no differences in IL-10 mRNA levels between any of the 3 melanoma cell lines tested and normal melanocytes. However, lower IL-10Rα expression was found in G361 and OCM-1 cells, and higher levels of IL-10Rβ were observed in G361 cells compared with normal melanocytes. GR-M cells did not exhibit any modifications in IL-10Rα and IL-10Rβ expression. miR-15a, miR-185, miR-211, and miR-30d were upregulated in G361 and OCM-1 cells, remaining at similar levels in GR-M cells. miR-409-3p and miR-605were down-regulated exclusively in G361 cells. Prediction tools revealed that miR-15a, miR-185, and miR-211 targeted IL-10Rα whereas none of the miRNAs exclusively downregulated in G361 cells targeted IL-10Rβ. Luciferase reporter and western blot assays showed that IL-10Rα expression is directly regulated by miR-15a, miR-185, and miR-211, either alone or in combination. An inverse expression pattern between IL-10Rα, on one side, and miR-15a, miR-185, and miR-211 on the other one was also shown in melanoma samples. Ectopic expression of individual miR-15a, miR-185, and miR-211, and even more their co-expression, caused a marked decrease in the proliferation rate of all the cell lines. Likewise, inhibition of any specific miRNA promoted cell growth, an effect that further increased when inhibition concerned all three miRNA. Moreover, specific knockdown of IL-10Rα prevented the proliferative effect of miRNA inhibitors.ConclusionsOur results support a key role of IL-10Rα in the development and progression of melanoma and suggest that the IL-10/IL-10 receptor system may become a new therapeutic target for melanoma treatment.

COXPRESdb: a database of comparative gene coexpression networks of eleven species for mammals

Coexpressed gene databases are valuable resources for identifying new gene functions or functional modules in metabolic pathways and signaling pathways. Although coexpressed gene databases are a fundamental platform in the field of plant biology, their use in animal studies is relatively limited. The COXPRESdb (http://coxpresdb.jp) provides coexpression relationships for multiple animal species, as comparisons of coexpressed gene lists can enhance the reliability of gene coexpression determinations. Here, we report the updates of the database, mainly focusing on the following two points. First, we updated our coexpression data by including recent microarray data for the previous seven species (human, mouse, rat, chicken, fly, zebrafish and nematode) and adding four new species (monkey, dog, budding yeast and fission yeast), along with a new human microarray platform. A reliability scoring function was also implemented, based on coexpression conservation to filter out coexpression with low reliability. Second, the network drawing function was updated, to implement automatic cluster analyses with enrichment analyses in Gene Ontology and in cis elements, along with interactive network analyses with Cytoscape Web. With these updates, COXPRESdb will become a more powerful tool for analyses of functional and regulatory networks of genes in a variety of animal species.

Long oligonucleotide microarrays for African green monkey gene expression profile analysis

Nonhuman primates, including African green monkey (AGM), are important models for biomedical research. The information on monkey genomes is still limited and no versatile gene expression screening tool is available. We tested human whole genome microarrays for cross-species reactivity with AGM transcripts using both long oligonucleotide arrays (60-mer probes) and short oligonucleotide arrays (25-mer). Using the long oligonucleotide arrays, we detected 4-fold more AGM transcripts than with the short oligonucleotide technology. The number of detected transcripts was comparable to that detected using human RNA, with 87% of the detected genes being shared between both species. The specificity of the signals obtained with the long oligonucleotide arrays was determined by analyzing the transcriptome of concanavalin A-activated CD4+ T cells vs. nonactivated T cells of two monkey species AGM and macaque. For both species, the genes showing the most significant changes in expression, such as IL-2R, were those known to be regulated in human CD4+ T cell activation. Finally, tissue specificity of the signals was established by comparing the transcription profiles of AGM brain and tonsil cells. In conclusion, the ABI human microarray platform provides a highly valuable tool for the assessment of AGM gene expression profiles.