1. ncRNA selection
After selection of the type of non-coding RNA of interest by clicking on the corresponding DNA structure, a text-box will be displayed. Here, one supplies the name of the ncRNA of which he/she wants to retrieve functional information. Upon entering the ncRNA name, a suggestion box will appear from which the user selects the appropriate ncRNA by clicking on it. Finally, by clicking on the "Next" button, a webpage will be displayed offering the user various options for retrieving relevant information concerning the supplied ncRNA.
2. Available ncRNA2function options
Depending on the type of ncRNA, different analysis options will be displayed. For long non-coding RNA molecules, only options for retrieving associated pathways are shown, while additional options for retrieving associated targets are available for miRNAs. Depending on the selected analysis, different customization options will be displayed. Each analysis option will be discussed in more detail in the paragraphs below.
2a. Associated pathways in selected datasets
When selecting the "Associated pathways in selected datasets" option - available for both miRNAs and lncRNAs - the user will be given the opportunity to choose the datasets and gene set for which he/she wants to retrieve gene set enrichment analysis (GSEA) results. Additional filtering options - to be performed on the results - include a gene set FDR value, an R cut-off and the possibility to display results associated with at least 2 or all of the selected datasets. Results can be displayed as a list or circos plot. In case a list visualization is selected, gene set hits will be ranked according to p-value, and the datasets in which each hit is present will be marked by a red (negative correlation) or green (positive correlation) dot. Linkouts to webpages containing information on a certain gene set are available by clicking on the corresponding gene set name. Another way to visualize the available GSEA results is by means of a circos plot. The data included on a circos plot is limited to the 200 most significantly correlated leading edge genes and their associated gene sets. For each gene set, the most significant leading edge genes along with their correlation value in each of the selected datasets are displayed by means of a heatmap. Arcs in the center of the plot connect leading edge genes in different gene sets and can be used to assess overlap between these sets. Finally, for miRNAs only, leading edge genes that are targets of the miRNA of interest are marked as well.
2b. Associated pathways across all datasets
This option will display all gene sets correlated with at least one of the available datasets for the ncRNA of interest. Results are visualized as a list, with the gene sets correlated with the most datasets on top, depicting the number of associated datasets. Clicking on a gene set name will open a table, showing more detailed information on the datasets such as their name, description and source, as well as the corresponding FDR and ES values.
2c. Associated targets in selected datasets
The "Associated targets in selected datasets" option behaves in much the same way as the first option (see paragraph 2a), but is only available for miRNAs. After selecting the datasets of interest and setting p and R filtering cut-offs, a circos plot is shown. The circos plot displays the significant correlated genes - that are targets of the miRNA of interest - and their correlation values in the different datasets.
2d. Associated targets across all datasets
The last option, also only available when selecting a miRNA, displays the same information as the previous option, however this time across all the available datasets. Results are shown as a list of miRNA target genes, associated with the number of datasets they are present in. Clicking on a gene name, will visualize additional information regarding the associated datasets such a name, description, source and p- and R-values.