GO enrichment
Philipp Bayer
01/10/2021
Last updated: 2021-12-20
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Knit directory: Amphibolis_Posidonia_Comparison/
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Lost in seagrasses
The GO enrichment does not work well on my laptop so I’m setting this to eval=FALSE and run it on a remote server. The script which writes the input files is a Python script in the scripts/ folder: findClustersUniqueToAquatic.py. This script parses Orthofinder output to pull out genome-specific groups and genes.
# give properly formatted background in format: GO:0005838 GSBRNA2T00088508001;GSBRNA2T00088313001;GSBRNA2T00035842001
#annAT <- readMappings('BACKGROUND.txt.gz', sep="\t", IDsep=";")
#save(annAT, file='annAtObject.RData')
load('annAtObject.RData')
allgenes <- unique(unlist(annAT))
compare <- function(genelistfile, outname, allgenes, annAT) {
# give file with your genes of interest, one gene_id per line
mygenes <-scan(genelistfile ,what="")
geneList <- factor(as.integer(allgenes %in% mygenes))
names(geneList) <- allgenes
GOdata <-new ("topGOdata", ontology = 'BP', allGenes = geneList, nodeSize = 5, annot=annFUN.GO2genes, GO2genes=annAT)
# using ClassicCount:
#test.stat <-new ("classicCount", testStatistic = GOFisherTest, name = "Fisher Test")
#resultsFisherC <-getSigGroups (GOdata, test.stat)
# using weight01:
weight01.fisher <- runTest(GOdata, statistic = "fisher")
# using ClassicCount:
# allRes <- GenTable(GOdata, classicFisher= resultsFisherC, topNodes = 30)
# using weight01:
allRes <- GenTable(GOdata, classicFisher=weight01.fisher,topNodes=30)#,topNodes=100)
names(allRes)[length(allRes)] <- "p.value"
p_values <- score(weight01.fisher)
adjusted_p <- p.adjust(p_values)
adjusted_p[adjusted_p < 0.05] %>% enframe() %>% write_csv('data/' + outname)
}lost in seagrasses, each one uniquely vs all terrestrials
compare('lost_in_amphi_vs_all.txt', 'missing_amphi_vs_all_GO.txt', allgenes, annAT)
compare('lost_in_posi_vs_all.txt', 'missing_posi_vs_all_GO.txt', allgenes, annAT)
compare('lost_in_zmar_vs_all.txt', 'missing_zmar_vs_all_GO.txt', allgenes, annAT)
compare('lost_in_zmuel_vs_all.txt', 'missing_zmuel_vs_all_GO.txt', allgenes, annAT)lost in seagrasses vs all terrestrials
Here we compare GO terms for seagrasses and aquatics (seagrasses+duckweeds) vs all terrestrials
compare('lost_in_seagrasses.txt', 'missing_seagrasses_GO.txt', allgenes, annAT)
compare('lost_in_aquatics.txt', 'missing_aquatics_GO.txt', allgenes, annAT)
compare('only_in_seagrasses.txt', 'only_seagrasses_GO.txt', allgenes, annAT)Seagrass only comparisons
Now we compare seagrasses within each other.
For the seagrass-only comparisons, I’m using a Seagrass-only background as that makes more biological sense to me.s
# give properly formatted background in format: GO:0005838 GSBRNA2T00088508001;GSBRNA2T00088313001;GSBRNA2T00035842001
#sannAT <- readMappings('SEAGRASSBACKGROUND.txt', sep="\t", IDsep=";")
#save(sannAT, file='sannAtObject.RData')
load('annAtObject.RData')
sallgenes <- unique(unlist(sannAT))
compare('lost_in_amphi.txt', 'lost_in_amphi_vs_seagrasses_GO.txt', sallgenes, sannAT)
compare('lost_in_posi.txt', 'lost_in_posi_vs_seagrasses_GO.txt', sallgenes, sannAT)
compare('lost_in_zmar.txt', 'lost_in_zmar_vs_seagrasses_GO.txt', sallgenes, sannAT)
compare('lost_in_zmuel.txt', 'lost_in_zmuel_vs_seagrasses_GO.txt', sallgenes, sannAT)Present in seagrasses
The opposite - which GO-terms are present only in one of the four species?
compare('only_in_amphi.txt', 'only_in_amphi_vs_seagrasses_GO.txt', sallgenes, sannAT)
compare('only_in_posi.txt', 'only_in_posi_vs_seagrasses_GO.txt', sallgenes, sannAT)
compare('only_in_zmar.txt', 'only_in_zmar_vs_seagrasses_GO.txt', sallgenes, sannAT)
compare('only_in_zmuel.txt', 'only_in_zmuel_vs_seagrasses_GO.txt', sallgenes, sannAT)Alright now we have all these different GO terms in all these files - we can send them to revigo for visualiation and some deduplication!
Revigo
This code is based on http://revigo.irb.hr/CodeExamples/revigo.R.txt
results_list <- list()
for (f in list.files('./data/', pattern='GO.txt')){
filename <- paste('./data/', f, sep='')
go_and_pvalues <- readChar(filename, file.info(filename)$size)
go_and_pvalues <- gsub(',', ' ', go_and_pvalues)
httr::POST(
url = "http://revigo.irb.hr/StartJob.aspx",
body = list(
cutoff = "0.7",
valueType = "pvalue",
# speciesTaxon = "4577", # zea mays
#speciesTaxon = '39947', # japonica
speciesTaxon = '3702', # arabidopsis
measure = "SIMREL",
goList = go_and_pvalues
),
# application/x-www-form-urlencoded
encode = "form"
) -> res
dat <- httr::content(res, encoding = "UTF-8")
jobid <- jsonlite::fromJSON(dat,bigint_as_char=TRUE)$jobid
# Check job status
running <- "1"
while (running != "0" ) {
httr::POST(
url = "http://revigo.irb.hr/QueryJobStatus.aspx",
query = list( jobid = jobid )
) -> res2
dat2 <- httr::content(res2, encoding = "UTF-8")
running <- jsonlite::fromJSON(dat2)$running
Sys.sleep(1)
}
# Fetch results
httr::POST(
url = "http://revigo.irb.hr/ExportJob.aspx",
query = list(
jobid = jobid,
namespace = "1",
type = "CSVTable"
)
) -> res3
dat3 <- httr::content(res3, encoding = "UTF-8")
dat3 <- stri_replace_all_fixed(dat3, "\r", "")
# Now we have a csv table in a string!
# read_csv does not like the ', ', it wants ','
dat <- read_csv(gsub(', ', ',', dat3), show_col_types = FALSE)
# do we even have results?
if(nrow(dat) == 0){next}
results_list[[f]] <- dat
}OK we have a list with all results in a big list. Now we can plot!
Making Supplementary Tables for the REVIGO results
Let’s also pull these terms out as tables
for( i in names(results_list)) {
print(i)
knitr::kable(results_list[[i]] %>% filter(Eliminated == FALSE), label=i)
}[1] "lost_in_amphi_GO.txt"
[1] "lost_in_posi_GO.txt"
[1] "lost_in_zmar_GO.txt"
[1] "lost_in_zmuel_GO.txt"
[1] "missing_amphi_vs_all_GO.txt"
[1] "missing_aquatics_GO.txt"
[1] "missing_arabidopsis_vs_all_GO.txt"
[1] "missing_posi_vs_all_GO.txt"
[1] "missing_seagrasses_GO.txt"
[1] "missing_zmar_vs_all_GO.txt"
[1] "missing_zmuel_vs_all_GO.txt"
[1] "only_in_posi_GO.txt"
[1] "only_in_zmar_GO.txt"
[1] "only_in_zmuel_GO.txt"
[1] "only_seagrasses_GO.txt"Venn diagram GO overlaps
Let’s also check which GO terms overlap between the 4 seagrasses!
Let’s have a look at all of these plots. Manually zooming in leads to ggrepel reloading labels, so on the small scale a lot of these plots don’t have labels.
for(i in names(plot_list)) {
file_name = paste('output/', i, '.png', sep='')
cowplot::save_plot(file_name, plot_list[[i]], base_height=8)
}How many shared lost GO-terms are there? Hopefully, all four species will have lost the most GO-terms.
a <- list(`P. australis` = results_list$missing_posi_vs_all_GO.txt$Name,
`A. antarctica` = results_list$missing_amphi_vs_all_GO.txt$Name,
`Z. marina` = results_list$missing_zmar_vs_all_GO.txt$Name,
`Z. muelleri` = results_list$missing_zmuel_vs_all_GO.txt$Name,
`A. thaliana` = results_list$missing_arabidopsis_vs_all_GO.txt$Name)
a_go_ids <- list(`P. australis` = results_list$missing_posi_vs_all_GO.txt$TermID,
`A. antarctica` = results_list$missing_amphi_vs_all_GO.txt$TermID,
`Z. marina` = results_list$missing_zmar_vs_all_GO.txt$TermID,
`Z. muelleri` = results_list$missing_zmuel_vs_all_GO.txt$TermID,
`A. thaliana` = results_list$missing_arabidopsis_vs_all_GO.txt$TermID)plot(euler(a),
quantities = TRUE,
fill = rev(wes_palette("Zissou1", 15, type = 'continuous')),
alpha = 0.5,
labels = list(font = 4))
upset(fromList(a), order.by='freq', )
a_no_ara <- list(`P. australis` = results_list$missing_posi_vs_all_GO.txt$Name,
`A. antarctica` = results_list$missing_amphi_vs_all_GO.txt$Name,
`Z. marina` = results_list$missing_zmar_vs_all_GO.txt$Name,
`Z. muelleri` = results_list$missing_zmuel_vs_all_GO.txt$Name)What are the shared GO-terms in seagrasses, WITHOUT the Ara loss??
setdiff(Reduce(intersect, a_no_ara), Reduce(intersect, a)) %>% enframe() %>% writexl::write_xlsx('./data/shared_lost_genes.xlsx')What if we remove Posidonia?
b <- list(`A. antarctica` = results_list$missing_amphi_vs_all_GO.txt$Name,
`Z. marina` = results_list$missing_zmar_vs_all_GO.txt$Name,
`Z. muelleri` = results_list$missing_zmuel_vs_all_GO.txt$Name)
intersections <- Reduce(intersect, b)
intersections[grepl('ethylene', intersections)][1] "jasmonic acid and ethylene-dependent systemic resistance,ethylene mediated signaling pathway"
[2] "regulation of ethylene-activated signaling pathway" OK we need a big list of all GO-terms here - which GO-term is lost in which species. That will be a supplementary table.
all_species <- c("P. australis","A. antarctica","Z. marina","Z. muelleri", 'A. thaliana')
all_go_terms <- Reduce(union, a)
all_go_ids <- Reduce(union, a_go_ids)
results_d <- data.frame('GOID' = character(),
'GO' = character(),
'P. australis' = character(),
'A. antarctica' = character(),
'Z. marina' = character(),
'Z. muelleri' = character(),
'A. thaliana' = character())
for (index in seq_along(all_go_terms)){
go <- all_go_terms[index]
go_id <- all_go_ids[index]
specs <- c()
for (species in names(a)) {
if ( length(a[[species]][grep(paste('^', go, '$', sep=''), a[[species]])]) > 0 ) {
specs <- c(specs, species)
}
}
results_d[index,] <- c(go_id, go, gsub('FALSE', 'Present', gsub('TRUE', 'Lost', all_species %in% specs)))
}writexl::write_xlsx(results_d, 'data/Lost_GO_terms_in_five_species.xlsx')after filtering for plant-specific GO-terms
We will use the GO-terms that are plant-specific as identified by the GOMAP paper. See https://github.com/wkpalan/GOMAP-maize-analysis/blob/main/6.plantSpecific/1.getSppSpecific.R or https://plantmethods.biomedcentral.com/articles/10.1186/s13007-021-00754-1
go_plant <- read_tsv('https://raw.githubusercontent.com/wkpalan/GOMAP-maize-analysis/main/data/go/speciesSpecificGOTerms.txt')Rows: 45031 Columns: 5-- Column specification --------------------------------------------------------
Delimiter: "\t"
chr (1): GOterm
dbl (4): NCBITaxon:10090, NCBITaxon:33090, NCBITaxon:3702, NCBITaxon:40674
i Use `spec()` to retrieve the full column specification for this data.
i Specify the column types or set `show_col_types = FALSE` to quiet this message.# taxon 33090 is Viridiplantae
plantSpecificGO <- go_plant %>% dplyr::filter(`NCBITaxon:33090`==1) %>% pull(GOterm)
plantSpecificGO <- c(plantSpecificGO,c("GO:0005575","GO:0008150","GO:0003674"))results_d %>% filter(GOID %in% plantSpecificGO) %>% writexl::write_xlsx('data/Lost_GO_terms_in_five_species.PlantSpecific.xlsx')Now let’s redo the Venn diagram with those filtered GO-terms
filters <- lapply(a_go_ids, function(ch) ch %in% plantSpecificGO)
newa <- list()
for (species in names(filters)) {
before <- a[[species]]
after <- before[filters[[species]]]
newa[[species]] <- after
}plot(euler(newa),
quantities = TRUE,
fill = rev(wes_palette("Zissou1", 15, type = 'continuous')),
alpha = 0.5,
labels = list(font = 4))
Not much difference?
sessionInfo()R version 4.1.0 (2021-05-18)
Platform: x86_64-w64-mingw32/x64 (64-bit)
Running under: Windows 10 x64 (build 19042)
Matrix products: default
locale:
[1] LC_COLLATE=English_Australia.1252 LC_CTYPE=English_Australia.1252
[3] LC_MONETARY=English_Australia.1252 LC_NUMERIC=C
[5] LC_TIME=English_Australia.1252
attached base packages:
[1] stats4 parallel stats graphics grDevices utils datasets
[8] methods base
other attached packages:
[1] rrvgo_1.4.4 UpSetR_1.4.0 eulerr_6.1.1
[4] ggrepel_0.9.1 stringi_1.7.5 httr_1.4.2
[7] wesanderson_0.3.6 rvest_1.0.2 forcats_0.5.1
[10] stringr_1.4.0 dplyr_1.0.7 purrr_0.3.4
[13] readr_2.0.2 tidyr_1.1.4 tibble_3.1.5
[16] ggplot2_3.3.5 tidyverse_1.3.1 topGO_2.44.0
[19] SparseM_1.81 GO.db_3.13.0 AnnotationDbi_1.54.1
[22] IRanges_2.26.0 S4Vectors_0.30.2 Biobase_2.52.0
[25] graph_1.70.0 BiocGenerics_0.38.0 workflowr_1.6.2
loaded via a namespace (and not attached):
[1] colorspace_2.0-2 ellipsis_0.3.2 rprojroot_2.0.2
[4] XVector_0.32.0 fs_1.5.0 rstudioapi_0.13
[7] farver_2.1.0 bit64_4.0.5 fansi_0.5.0
[10] lubridate_1.8.0 xml2_1.3.2 cachem_1.0.6
[13] GOSemSim_2.18.1 knitr_1.36 polyclip_1.10-0
[16] jsonlite_1.7.2 broom_0.7.9 gridBase_0.4-7
[19] dbplyr_2.1.1 png_0.1-7 pheatmap_1.0.12
[22] shiny_1.7.1 compiler_4.1.0 backports_1.2.1
[25] assertthat_0.2.1 fastmap_1.1.0 cli_3.0.1
[28] later_1.3.0 htmltools_0.5.2 tools_4.1.0
[31] igraph_1.2.7 NLP_0.2-1 gtable_0.3.0
[34] glue_1.4.2 GenomeInfoDbData_1.2.6 Rcpp_1.0.7
[37] slam_0.1-48 cellranger_1.1.0 jquerylib_0.1.4
[40] vctrs_0.3.8 Biostrings_2.60.2 writexl_1.4.0
[43] polylabelr_0.2.0 xfun_0.27 mime_0.12
[46] lifecycle_1.0.1 zlibbioc_1.38.0 scales_1.1.1
[49] treemap_2.4-3 vroom_1.5.5 hms_1.1.1
[52] promises_1.2.0.1 RColorBrewer_1.1-2 yaml_2.2.1
[55] curl_4.3.2 memoise_2.0.0 gridExtra_2.3
[58] sass_0.4.0 RSQLite_2.2.8 highr_0.9
[61] GenomeInfoDb_1.28.4 rlang_0.4.12 pkgconfig_2.0.3
[64] matrixStats_0.61.0 bitops_1.0-7 evaluate_0.14
[67] lattice_0.20-44 labeling_0.4.2 cowplot_1.1.1
[70] bit_4.0.4 tidyselect_1.1.1 plyr_1.8.6
[73] magrittr_2.0.1 R6_2.5.1 generics_0.1.1
[76] DBI_1.1.1 pillar_1.6.4 haven_2.4.3
[79] whisker_0.4 withr_2.4.2 KEGGREST_1.32.0
[82] RCurl_1.98-1.5 modelr_0.1.8 crayon_1.4.1
[85] wordcloud_2.6 utf8_1.2.2 tzdb_0.1.2
[88] rmarkdown_2.11 grid_4.1.0 readxl_1.3.1
[91] data.table_1.14.2 blob_1.2.2 git2r_0.28.0
[94] reprex_2.0.1 digest_0.6.28 xtable_1.8-4
[97] tm_0.7-8 httpuv_1.6.3 munsell_0.5.0
[100] bslib_0.3.1