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Use Case #1: Functional Annotation of Genes Sharing a Common Evolutionary History

Evolutionary Transcriptomics aims to predict stages or periods of evolutionary conservation in biological processes on the transcriptome level. However, finding genes sharing a common evolutionary history could reveal how the the biological process might have evolved in the first place.

In this Use Case we will combine functional and biological annotation obtained with biomartr with enriched genes obtained with PlotEnrichment().

Step 1

For the following example we will use the dataset an enrichment analyses found in PlotEnrichment().

Install and load the myTAI package:

# install myTAI
install.packages("myTAI")

# load myTAI
library(myTAI)

Download the Phylostratigraphic Map of D. rerio:

# download the Phylostratigraphic Map of Danio rerio
# from Sestak and Domazet-Loso, 2015

The dataset comes from Supplementary file 3 of this publication: https://academic.oup.com/mbe/article/32/2/299/1058654#77837069

After downloading Supplementary file 3, you will find the file TableS3-2.xlsx which can be used in the following biomartr functions.

Read the *.xlsx file storing the Phylostratigraphic Map of D. rerio and format it for the use with myTAI:

# install the readxl package
install.packages("readxl")

# load package readxl
library(readxl)

# read the excel file
DrerioPhyloMap.MBEa <- read_excel("TableS3-2.xlsx", sheet = 1, skip = 4)

# format Phylostratigraphic Map for use with myTAI
Drerio.PhyloMap <- DrerioPhyloMap.MBEa[ , 1:2]

# have a look at the final format
head(Drerio.PhyloMap)
  Phylostrata            ZFIN_ID
1           1 ZDB-GENE-000208-13
2           1 ZDB-GENE-000208-17
3           1 ZDB-GENE-000208-18
4           1 ZDB-GENE-000208-23
5           1  ZDB-GENE-000209-3
6           1  ZDB-GENE-000209-4

Now, Drerio.PhyloMap stores the Phylostratigraphic Map of D. rerio which is used as background set to perform enrichment analyses with PlotEnrichment() from myTAI.

Enrichment Analyses

Now, the PlotEnrichment() function visualizes the over- and underrepresented Phylostrata of brain specific genes when compared with the total number of genes stored in the Phylostratigraphic Map of D. rerio.

library(readxl)

# read expression data (organ specific genes) from Sestak and Domazet-Loso, 2015
Drerio.OrganSpecificExpression <- read_excel("TableS3-2.xlsx", sheet = 2, skip = 3)

# select only brain specific genes
Drerio.Brain.Genes <- unlist(unique(na.omit(Drerio.OrganSpecificExpression[ , "brain"])))

# visualize enriched Phylostrata of genes annotated as brain specific
PlotEnrichment(Drerio.PhyloMap,
               test.set     = Drerio.Brain.Genes,
               measure      = "foldchange",
               use.only.map = TRUE,
               legendName   = "PS")

Users will observe that for example brain genes deriving from PS5 are significantly enriched.

Now we can select all brain genes originating in PS5 using the SelectGeneSet() function from myTAI. Please notice that SelectGeneSet() can only be used with phylostratigraphic maps only (use.map.only = TRUE argument) since myTAI version > 0.3.0.

BrainGenes <- SelectGeneSet(ExpressionSet = Drerio.PhyloMap,
                            gene.set      = Drerio.Brain.Genes,
                            use.only.map  = TRUE)

# select only brain genes originating in PS5
BrainGenes.PS5 <- BrainGenes[which(BrainGenes[ , "Phylostrata"] == 5), ]

# look at the results
head(BrainGenes.PS5)
      Phylostrata           ZFIN_ID
14851           5 ZDB-GENE-000210-6
14852           5 ZDB-GENE-000210-7
14853           5 ZDB-GENE-000328-4
14856           5 ZDB-GENE-000411-1
14857           5 ZDB-GENE-000427-4
14860           5 ZDB-GENE-000526-1

Now users can perform the biomart() function to obtain the functional annotation of brain genes originating in PS5.

For this purpose, first we need to find the filter name of the corresponding gene ids such as ZDB-GENE-000210-6.

# find filter for zfin.org ids
organismFilters("Danio rerio", topic = "zfin_id")
                            name                           description             dataset
52                  with_zfin_id                       with ZFIN ID(s) drerio_gene_ensembl
53  with_zfin_id_transcript_name          with ZFIN transcript name(s) drerio_gene_ensembl
103                      zfin_id ZFIN ID(s) [e.g. ZDB-GENE-060825-136] drerio_gene_ensembl
274                 with_zfin_id                       with ZFIN ID(s)    drerio_gene_vega
286                      zfin_id ZFIN ID(s) [e.g. ZDB-GENE-121214-212]    drerio_gene_vega
366                 with_zfin_id                       with ZFIN ID(s) drerio_gene_ensembl
367 with_zfin_id_transcript_name          with ZFIN transcript name(s) drerio_gene_ensembl
417                      zfin_id ZFIN ID(s) [e.g. ZDB-GENE-060825-136] drerio_gene_ensembl
588                 with_zfin_id                       with ZFIN ID(s)    drerio_gene_vega
600                      zfin_id ZFIN ID(s) [e.g. ZDB-GENE-121214-212]    drerio_gene_vega
680                 with_zfin_id                       with ZFIN ID(s) drerio_gene_ensembl
681 with_zfin_id_transcript_name          with ZFIN transcript name(s) drerio_gene_ensembl
731                      zfin_id ZFIN ID(s) [e.g. ZDB-GENE-060825-136] drerio_gene_ensembl
902                 with_zfin_id                       with ZFIN ID(s)    drerio_gene_vega
914                      zfin_id ZFIN ID(s) [e.g. ZDB-GENE-121214-212]    drerio_gene_vega
                    mart
52  ENSEMBL_MART_ENSEMBL
53  ENSEMBL_MART_ENSEMBL
103 ENSEMBL_MART_ENSEMBL
274 ENSEMBL_MART_ENSEMBL
286 ENSEMBL_MART_ENSEMBL
366 ENSEMBL_MART_ENSEMBL
367 ENSEMBL_MART_ENSEMBL
417 ENSEMBL_MART_ENSEMBL
588 ENSEMBL_MART_ENSEMBL
600 ENSEMBL_MART_ENSEMBL
680 ENSEMBL_MART_ENSEMBL
681 ENSEMBL_MART_ENSEMBL
731 ENSEMBL_MART_ENSEMBL
902 ENSEMBL_MART_ENSEMBL
914 ENSEMBL_MART_ENSEMBL

Now users can retrieve the corresponding GO attribute of D. rerio with organismAttributes.

# find go attribute term for D. rerio
organismAttributes("Danio rerio", topic = "go")
                                              name                             description
33                                           go_id                       GO Term Accession
36                                 go_linkage_type                   GO Term Evidence Code
38                            goslim_goa_accession                 GOSlim GOA Accession(s)
39                          goslim_goa_description                  GOSlim GOA Description
516                  ggorilla_homolog_ensembl_gene                 Gorilla Ensembl Gene ID
517  ggorilla_homolog_canomical_transcript_protein      Canonical Protein or Transcript ID
518               ggorilla_homolog_ensembl_peptide              Gorilla Ensembl Protein ID
519                    ggorilla_homolog_chromosome                 Gorilla Chromosome Name
520                   ggorilla_homolog_chrom_start           Gorilla Chromosome Start (bp)
521                     ggorilla_homolog_chrom_end             Gorilla Chromosome End (bp)
522                ggorilla_homolog_orthology_type                           Homology Type
523                       ggorilla_homolog_subtype                                Ancestor
524          ggorilla_homolog_orthology_confidence    Orthology confidence [0 low, 1 high]
525                       ggorilla_homolog_perc_id   % Identity with respect to query gene
526                    ggorilla_homolog_perc_id_r1 % Identity with respect to Gorilla gene
527                            ggorilla_homolog_dn                                      dN
528                            ggorilla_homolog_ds                                      dS
1240                                         go_id                                   GO ID
1241                                      quick_go                             Quick GO ID
1370                                         go_id                       GO Term Accession
1373                               go_linkage_type                   GO Term Evidence Code
1375                          goslim_goa_accession                 GOSlim GOA Accession(s)
1376                        goslim_goa_description                  GOSlim GOA Description
1853                 ggorilla_homolog_ensembl_gene                 Gorilla Ensembl Gene ID
1854 ggorilla_homolog_canomical_transcript_protein      Canonical Protein or Transcript ID
1855              ggorilla_homolog_ensembl_peptide              Gorilla Ensembl Protein ID
1856                   ggorilla_homolog_chromosome                 Gorilla Chromosome Name
1857                  ggorilla_homolog_chrom_start           Gorilla Chromosome Start (bp)
1858                    ggorilla_homolog_chrom_end             Gorilla Chromosome End (bp)
1859               ggorilla_homolog_orthology_type                           Homology Type
1860                      ggorilla_homolog_subtype                                Ancestor
1861         ggorilla_homolog_orthology_confidence    Orthology confidence [0 low, 1 high]
1862                      ggorilla_homolog_perc_id   % Identity with respect to query gene
1863                   ggorilla_homolog_perc_id_r1 % Identity with respect to Gorilla gene
1864                           ggorilla_homolog_dn                                      dN
1865                           ggorilla_homolog_ds                                      dS
2577                                         go_id                                   GO ID
2578                                      quick_go                             Quick GO ID
2707                                         go_id                       GO Term Accession
2710                               go_linkage_type                   GO Term Evidence Code
2712                          goslim_goa_accession                 GOSlim GOA Accession(s)
2713                        goslim_goa_description                  GOSlim GOA Description
3190                 ggorilla_homolog_ensembl_gene                 Gorilla Ensembl Gene ID
3191 ggorilla_homolog_canomical_transcript_protein      Canonical Protein or Transcript ID
3192              ggorilla_homolog_ensembl_peptide              Gorilla Ensembl Protein ID
3193                   ggorilla_homolog_chromosome                 Gorilla Chromosome Name
3194                  ggorilla_homolog_chrom_start           Gorilla Chromosome Start (bp)
3195                    ggorilla_homolog_chrom_end             Gorilla Chromosome End (bp)
3196               ggorilla_homolog_orthology_type                           Homology Type
3197                      ggorilla_homolog_subtype                                Ancestor
3198         ggorilla_homolog_orthology_confidence    Orthology confidence [0 low, 1 high]
3199                      ggorilla_homolog_perc_id   % Identity with respect to query gene
3200                   ggorilla_homolog_perc_id_r1 % Identity with respect to Gorilla gene
3201                           ggorilla_homolog_dn                                      dN
3202                           ggorilla_homolog_ds                                      dS
3914                                         go_id                                   GO ID
3915                                      quick_go                             Quick GO ID
                 dataset                 mart
33   drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
36   drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
38   drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
39   drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
516  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
517  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
518  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
519  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
520  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
521  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
522  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
523  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
524  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
525  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
526  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
527  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
528  drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1240    drerio_gene_vega ENSEMBL_MART_ENSEMBL
1241    drerio_gene_vega ENSEMBL_MART_ENSEMBL
1370 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1373 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1375 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1376 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1853 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1854 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1855 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1856 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1857 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1858 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1859 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1860 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1861 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1862 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1863 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1864 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
1865 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
2577    drerio_gene_vega ENSEMBL_MART_ENSEMBL
2578    drerio_gene_vega ENSEMBL_MART_ENSEMBL
2707 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
2710 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
2712 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
2713 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3190 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3191 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3192 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3193 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3194 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3195 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3196 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3197 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3198 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3199 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3200 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3201 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3202 drerio_gene_ensembl ENSEMBL_MART_ENSEMBL
3914    drerio_gene_vega ENSEMBL_MART_ENSEMBL
3915    drerio_gene_vega ENSEMBL_MART_ENSEMBL

Now users can specify the filter zfin_id and attribute go_id to retrieve the GO terms of corresponding gene ids (Please note that this will take some time).

# retrieve GO terms of D. rerio brain genes originating in PS5
GO_tbl.BrainGenes <- biomart(genes      = unlist(BrainGenes.PS5[ , "ZFIN_ID"]),
                             mart       = "ENSEMBL_MART_ENSEMBL",
                             dataset    = "drerio_gene_ensembl",
                             attributes = "go_id",
                             filters    = "zfin_id")

head(GO_tbl.BrainGenes)
            zfin_id      go_id
1 ZDB-GENE-000210-6 GO:0060037
2 ZDB-GENE-000210-6 GO:0046983
3 ZDB-GENE-000210-7 GO:0046983
4 ZDB-GENE-000328-4 GO:0007275
5 ZDB-GENE-000328-4 GO:0007166
6 ZDB-GENE-000328-4 GO:0035567