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Basic Navigation

Overview

RiboVision 2 is operated via the Main Navigation panel for selection/filtering and contains three main applets:

  • Alignment Viewer for representation of multiple sequence alignments.
  • RNA Topology Viewer for depiction of RNA secondary structure layouts.
  • Mol* viewer for visualization of three-dimensional structures.

The applets appear in a consecutive manner upon selecting appropriate data.

Side Bar The main Navigation panel is designed in the interactive fashion and enables the users to selects or upload enables various objects sequentially. In RiboVision mode the Navigation Pannel contains a taxonomic browser, which enables the recursive selection of species from one or multiple taxonomic groups. RiboVision 2 contains preloaded alignments for 16S/18S, 23S/28S, 5.8S, and 5S rRNAs. RiboVision 2 calculates gap-adjusted nucleotide frequencies from these alignments. These frequencies are then utilized to compute phylogenetic conservation scores, employing methods such as Shannon Entropy or TwinCons (described below).
After loading the multiple sequence alignment (MSA), the corresponding RNA structure must be chosen. This is accomplished by either typing in or selecting a four-character PDB ID from the list of available ribosomal structures that appear in the Main Navigation panel. This list is dynamically generated via an API from ribosome.xyz, ensuring real-time updates of the available ribosomal structures. The server then filters and displays all accessible RNA chains within the provided PDB complex. It becomes the user's responsibility to select a chain that aligns with the MSA. Once the chain is selected, it will be visualized in the 3D Mol* viewer. Only a single RNA chain is shown by default. The users also have an option to display the entire complex by checking the box “Show ribosomal context in 3D".
The server will automatically process the computations of contacts between the selected RNA chain and ribosomal proteins in the selected complex at the backend. The results of computations will appear in the Main Navigation Panel. Similarly, chemical modifications of the selected RNA chain will be listed in the Main Navigation Panel, if their annotations are available in the corresponding ribosomal cif file. In order to view these data in 2D and 3D representations, the users should select the desired protein contacts of chemical modifications by checking the corresponding boxes (or select all of them) and press 'Submit Proteins' or 'Submit modifications 'button to trigger their visualization in the applets. The RNA nucleotides in the 2D and 3D applet will be highlighted by different colors according to the data supplied. When proteins are selected and submitted, their 3D structures will also appear in the 3D viewer. The proteins will appear iteratively, it may take several seconds for to update the entire state. The 3D representations of proteins will be colored individually matching the colors of the corresponding protein-RNA contacts.

Selecting phylogenetic group(s)

To retrieve a specified alignment from the DESIRE database, follow the steps below:

  1. Click on the dropdown menu Select a phylogenetic group; the three dropdown nodes are labeled with the three domains of life: Eukarya, Bacteria, and Archaea.
  2. Click on the left-side drop-down arrow of any phylogenetic group in the dropdown menu; the phylogenetic subgroups of that group become visible. For example, upon clicking the left-side arrow of “Bacteria,” the following groups are made visible to the user: “Bacteroidetes,” “Chlorobi,” “Cyanobacteria,” “Proteobacteria,” etc. This step is recursive.
  3. Add a phylogenetic group to the list of phylogenetic groups for which the alignment will be retrieved. This list is displayed at the top of the drop-down structure. Note that when zero groups are selected this field displays the text Select a phylogenetic group. Multiple groups can be selected.
  4. The user can also directly search for a phylogenetic group by typing in its name.

Selecting an RNA alignment

Upon selection of a set of phylogenetic groups (See Selecting phylogenetic group(s)), a list of molecules (LSU or SSU) appears, and once a desired group is selected, compatible RNA alignments are pulled from the RiboVision 2 database in a new dropdown menu. The alignments list is filtered to show alignments that contain all selected phylogenetic groups. For example, if a user selects Archaea, Bacteria and Eukarya, only universal RNA alignments will be shown. Once the user selects an RNA alignment from the available ones, the alignment is displayed in the alignment viewer; the rows of the alignment are phylogenetic groups, and the columns of the alignment are arranged according to individual amino acid indices.

Selecting a structure for visualization and mapping

Once an alignment of a specified RNA is selected, the option to select its structure for visualization and mapping becomes available. Selecting a structure is a two-step process:

  1. First the user selects a 3D structure available from the RCSB/PDBe. Structures from the PDBe are filtered by the polymer of the selected alignment via the RiboXYZ.org API. Filtered PDB IDs are available in a dropdown menu when using the DESIRE database. For user convenience, the first three PDB ID options are never filtered and are those for the cytosolic ribosomes of E. coli (7K00), P. furiosus (4V6U), and H. sapiens (4UG0).
  2. Once a PDB ID is selected, the available RNA chains of the selected structure are filtered and shown in the next window. The user needs to select one chain that has to match the polymer in the alignment viewer. The selected chain will load the topology and Mol* viewers.

Creating a structure-alignment mapping

To ensure a match between the selected MSA and the sequence from a selected structure and to properly visualize the data, we compute an alignment between the sequences of the structure and the alignment. This is done internally on the server using the mafft program with the –addfull option (Katoh; 10.1093/molbev/mst010) and it does not require any action from the user. However, if the structural sequence has extra positions compared to the alignment, RiboVision 2 displays an error message located between the alignment and 2D/3D viewers. The message warns the user how many positions failed to map properly:
Warning, poor alignment between selected MSA and structure!!!
This message appears when the selected structure is mismatched with the pre-selected/pre-uploaded MSA in a range over 50 nts. This may happen due to:

  1. improper selection of an MSA/structure pair (e.g. 23S rRNA chain is selected in the structure and mistakenly mapped onto 16S rRNA MSA) or
  2. large discrepancy in the selected MSA and 3D structure (e.g. in case of selecting an MSA alignment for bacteria and 3D structure for a mammalian species).

In both cases the warning message is aimed to draw the attention of the user.

Selecting attribute data for mapping

The user has an option to map either calculated mapping data (see Advanced features) or custom data (supplied by a user) onto the 2D and 3D viewers. Available data attributes may be selected from a dropdown menu in the lower right corner of the RNA2D viewer. There is also an option in the sidebar to upload custom mapping data. This data should be uploaded as a .csv; an example file is provided on the RiboVision 2 site. Once the custom data has been uploaded, it will be added as a mapping option in the dropdown menu of the RNA2D viewer applet.