⊟AureoWiki - The Wiki way of Staphylococcus aureus annotation

AureoWiki is the first tool to support curated annotation of S. aureus biological entities in a Wiki based manner. At the currently available state, AureoWiki provides information for an S. aureus pan-genome of 33 strains. Furthermore, detailed information on the genes and proteins of S. aureus strains COL, NCTC8325, N315, Newman and USA300_FPR3757 is provided. These five strains were selected because of their detailed annotation (N315) or their clinical or scientific importance. The AureoWiki gene pages contain a lot of information from a variety of sources, including e.g. data on protein function and localization, transcriptional regulation, and gene expression. Orthologous genes of the individual strains are linked by a pan-genome gene identifier and a unified gene name. The main objective of AureoWiki is to facilitate the transfer of knowledge gained in studies with different S. aureus strains, thus supporting functional annotation and better understanding of this organism. Some direct links may give an impression of typical gene / protein pages: hld, eno, hlY, splA, pflB, saeR, sarA

The scientific community is invited to extend the current information. This can be performed via the Edit icon on the top right of each page or the [edit] links on top of each section or inside some sections. Changes can be made as a registered user (you can setup your own account by using the button with the small down arrow next to the Log in link) or by answering a Staphylococcus research field associated expert question before the saving process is initiated. Any user will easily be able to add information to the AureoWiki as described here.

Information preceded by a filled bullet point is fed from a database behind the AureoWiki and cannot be changed. This is because we propose that especially sequence based information should be consistently maintained in accordance with the RefSeq annotation and other databases. An [edit] link makes it possible to add user generated content or comments. User added information is indicated by open bullets. At some places we added placeholders to ask for user input (e.g. phenotype section of the strain-specific gene pages) to complement the currently available information.

The Main Page presents an easy to use Search mask. Here the user can search for locus tags, gene symbols or keywords. The [Getting Started] button brings you to the actual page.

The bar of register tabs on top of the AureoWiki pages allows switching between the pan-genome pages and the strain-specific pages for a gene. The pages are linked via an orthologue reference table that was constructed within our work and can be downloaded using the [Downloads] button. What kind of information you can find in the AureoWiki and where this information comes from will be explained in the following chapters:

⊟The S.aureus Pan Genome and the AureoWiki Pan Genome Pages

For a comparative analysis of S. aureus strains an overall alignment of a publicly available set of 33 strain-specific S. aureus genomes from NCBI RefSeq was performed.

NC_002951	COL	NC_002745	N315	NC_007795	NCTC_8325	NC_009641	Newman
NC_007793	USA300_FPR3757
NC_017340	04_02981	NC_013450	ED98	NC_016928	M013	NC_007622	RF122
NC_018608	08BA02176	NC_017763	HO_5096_0412	NC_002952	MRSA252	NC_017333	ST398
NC_017351	11819_97	NC_009632	JH1	NC_016941	MSHR1132	NC_017347	T0131
NC_022222	6850	NC_009487	JH9	NC_002953	MSSA476	NC_017342	TCH60
NC_017673	71193	NC_017341	JKD6008	NC_009782	Mu3	NC_017331	TW20
NC_017343	ECT_R 2	NC_017338	JKD6159	NC_002758	Mu50	NC_010079	USA300_TCH1516
NC_017337	ED133	NC_017349	LGA251	NC_003923	MW2	NC_016912	VC40

On this basis it became possible to construct a positionally corrected and all genes containing pan-genome. This was used to assign homologous genes (at least 50% identity on DNA and 70% identity on protein level) of different S. aureus strains to orthologous gene groups, so called unified pan-genome gene IDs. The corresponding pan-genome gene symbols are species-wide unified and unique gene name resulting from a manual curation effort. In brief, pan gene symbols were assigned based on strain-specific gene symbols from NCBI-RefSeq and/or supported by relevant publications or, alternatively, the name of the orthologous Bacillus subtilis gene if it is not a “y”-name.

The pan-genome was determined by the group of Kay Nieselt (Center for Bioinformatics, University of Tübingen) by progressiveMAUVE based total genome alignment followed by an iterative refinement of orthologue groups with special attention to gene synteny by using OrthologyPredator and PanGee (L. Backert, A. Henning, unpublished data).

The so called pan-genome pages, which can be accessed by using the most left register tab on top of the AureoWiki pages, present the following information:

A unique pan-genome gene ID (pan ID) resulting from the order of all genes within the S. aureus pan-genome, followed by the pan-genome gene symbol and a list of gene descriptions that were extracted from the RefSeq annotations of all 33 S. aureus genomes. Next, you find the strand and the start and end positions of the pan-genome genes within the positionally standardized pan-genome as well as the so called synteny blocks, which define phylogenetically conserved genome regions. How often a gene has orthologues genes in the 33 S. aureus genomes used for the pan-genome construction is shown in the occurrence entry.

In the following Orthologs section, all 33 S. aureus strains are listed in a fixed order and, if the gene is present in the respective strain, the locus tag is shown together with the strain-specific gene name, if assigned, from the RefSeq annotation.

A graphical display of the respective genomic region of the N315, COL, NCTC8325, Newman and USA300_FPR3757 strains highlights phylogenetic conservation. The used colors encode for gene functional assignments (see table below) performed by using a collection of Hidden Markov models of TIGRFAMs.

Meta Function	Gene Functional Class (TIGRFam Main Role)	Color Code
Envelope	Cell envelope
Cellular processes	Cellular processes
Metabolism	Amino acid biosynthesis
	Biosynthesis of cofactors, prosthetic groups, and carriers
	Central intermediary metabolism
	Energy metabolism
	Fatty acid and phospholipid metabolism
	Purines, pyrimidines, nucleosides, and nucleotides
	Transport and binding proteins
Genetic Info processing	DNA metabolism
	Mobile and extrachromosomal element functions
	Protein fate
	Protein synthesis
	Transcription
Regulation	Regulatory functions
	Signal transduction
Unknown function	Hypothetical proteins
	Unknown function
RNAs	RNA genes

⊟The Strain Specific Pages

The Summary section at the top of each page contains the locus tag, the gene name and function of the gene product from the RefSeq annotation as well as the pan locus tag and the pan gene symbol. In the following Genome View (based on a vectorized (SVG) file format), condensed genome information is provided, initially aligned to the position of the respective gene. The genome position in the genome browser can be changed by dragging the slider. By clicking on gene arrows, the user is transferred to the corresponding gene page, thus enabling a page by page walking through the genome. Colors correspond to the gene functional categories as described above. Finally, the genome browser combines for each strain the well-established RefSeq annotation and the new RefSeq annotation introduced in 2015 , thus directly showing the differences in gene content and/or coordinates.

The Gene section contains the information about the gene. It covers basic information as in the Summary section, complemented by the gene coordinates, gene length, essentiality, DNA sequence, and external accession numbers with links to the gene-specific database entries.

The largest section of the page, the Protein section, is devoted to the encoded protein. It shows, amongst others, the protein length, the molecular weight and isoelectric point, catalyzed reaction, protein function assignments (see next paragraph), protein interaction partners, and subcellular localization. Finally, the Protein section contains database links (NCBI Protein database and UniProt), the protein sequence and experimental data including protein localization and absolute quantification of cytoplasmic proteins .

Functional assignments have been generated as follows:

1. For enzymes the catalytic activity is provided by the EC number (extracted from NCBI RefSeq and UniProt databases), complemented by the corresponding enzyme name and reaction equation (extracted from ExPASy).
2. The assignment of protein sequences to TIGRFAMs protein families is based on TIGRFAM Hidden Markov Models (HMM) using hmmscan of the HMMER3 software package . The display of TIGRFAMs is ordered according to their HMM scores as a significance measure of the assignment and possesses a tree like structure including the TIGR role categories (main role and sub role) and an added meta level summarizing the TIGR main roles (see also color table above).
3. As described for TIGRFAMs, the assignment of sequences to Pfam protein families is based on HMMs and uses the HMMER package. Pfams with the highest HMM scores are shown first. A large part of Pfams is grouped into clans (evolutionary related families), which are displayed on top of the Pfam annotation.
4. Assignment of predicted protein functions is obtained from the SEED database , a comparative genomics database based on expert annotation of subsystems (sets of related functional roles). By default the lists of assigned predicted functions are collapsed and show only the hit with the highest score, but can be expanded by clicking the plus sign.

The following section of the gene page provides information on Expression & Regulation, including the predicted operon structure obtained from MicrobesOnline , gene expression regulation as well as gene expression profiles. Data on transcription factor regulons, alternative RNA polymerase sigma factors (SigB and SigH) and regulatory RNAs was retrieved from the RegPrecise database and published literature. Gene expression data from two large-scale studies covering a wide range of growth, stress and infection-related conditions were included in AureoWiki by graphical representations of condition-dependent mRNA levels and protein induction profiles on the gene pages of strains NCTC 8325 and COL, respectively.

All data are provided with links to the external data sources, including various databases and published literature. References are indicated by the book symbol. Details of the corresponding publication are displayed by mouse-over. A list of additional literature is found under "Relevant publications" at the bottom of the page.