Jiorgos Kourelis 30 July, 2021
refplantnlR is a package for drawing NLR domain architecture. Either
provide it with the NLRtracker output, or load the RefPlantNLR dataset
or NCBI RefSeq NLR dataset and the function does (most of) the rest!
To install you’ll need the devtools R package. From the R console in
RStudio type
install.packages("devtools")Once devtools is installed you can use that to install refplantnlR
devtools::install_github("JKourelis/refplantnlR")The first step is to load our domain architecture data file and do some basic filtering
First read in your domain file output from NLRtracker. You can do that
with read.delim(). Must contain a minimum of one “CHAIN” as data$type.
library(dplyr)
library(refplantnlR)
RefPlantNLR <- RefPlantNLR
head(RefPlantNLR)
#> Status seqname Subclass Domain_simplified Species Order
#> 1 NLR 3gG2 CC-NLR CNL Glycine max Fabales
#> 2 NLR Adnr1 CC-NLR OCNLO Triticum aestivum Poales
#> 3 NLR Adnr1-RGA4 CC-NLR CNL Triticum aestivum Poales
#> 4 NLR ADR1 CCR-NLR RNL Arabidopsis thaliana Brassicales
#> 5 NLR ADR1-L1 CCR-NLR RNL Arabidopsis thaliana Brassicales
#> 6 NLR ADR1-L2 CCR-NLR RNL Arabidopsis thaliana Brassicales
#> Pathogen Genus Representative type description start end
#> 1 Virus Glycine YES CHAIN chain 1 1129
#> 2 Arthropods Triticum YES CHAIN chain 1 1482
#> 3 HELPER Triticum NO CHAIN chain 1 997
#> 4 HELPER Arabidopsis YES CHAIN chain 1 787
#> 5 HELPER Arabidopsis YES CHAIN chain 1 816
#> 6 HELPER Arabidopsis YES CHAIN chain 1 811We can now filter the NLRs to select only the ones with e.g integrated
domains, using filter()
filtered_RefPlantNLR <- filter(RefPlantNLR, grepl("O", Domain_simplified),
Status == "NLR")
head(filtered_RefPlantNLR)
#> Status seqname Subclass Domain_simplified Species
#> 1 NLR Adnr1 CC-NLR OCNLO Triticum aestivum
#> 2 NLR CHS3 TIR-NLR TNLO Arabidopsis thaliana
#> 3 NLR DAR5 CCR-NLR RNOO Arabidopsis thaliana
#> 4 NLR Pi-ta CC-NLR CNLO Oryza sativa Japonica Group
#> 5 NLR Pi1-5 CC-NLR CONL Oryza sativa Indica Group
#> 6 NLR Pi5-3 CC-NLR CNLO Oryza sativa Japonica Group
#> Order Pathogen Genus Representative type description start
#> 1 Poales Arthropods Triticum YES CHAIN chain 1
#> 2 Brassicales AUTOIMMUNITY Arabidopsis YES CHAIN chain 1
#> 3 Brassicales <NA> Arabidopsis YES CHAIN chain 1
#> 4 Poales Fungi Oryza YES CHAIN chain 1
#> 5 Poales Fungi Oryza YES CHAIN chain 1
#> 6 Poales <NA> Oryza YES CHAIN chain 1
#> end
#> 1 1482
#> 2 1613
#> 3 702
#> 4 1033
#> 5 1143
#> 6 1049We can now draw the domain architecture
p <- draw_nlr(filtered_RefPlantNLR)
p
We can modify the output to also plot the NB-ARC motifs
filtered_RefPlantNLR <- filter(RefPlantNLR, Subclass == "CCR-NLR")
p <- draw_nlr(filtered_RefPlantNLR, label_motifs = TRUE)
p
Finally, specifically for the RefPlantNLR dataset we can also label the individual LRRs
filtered_RefPlantNLR <- filter(RefPlantNLR, Subclass == "CCG10-NLR")
p <- draw_nlr(filtered_RefPlantNLR, label_repeats = TRUE)
p
With the prepackaged NLRtracker output from all plant NCBI RefSeq proteomes we can do some additional analysis besides plotting the domain architecture. For example we can plot the number of NLR loci that contain potential integratd domains
library(tidyverse)
# The dataset
RefSeq_NLR <- RefSeq_NLR
## Filtering to select a single splice-variant per locus, prioritizing the ones with integrated domains
RefSeq_NLR_LOCUS <- RefSeq_NLR %>%
filter(Status %in% c("NLR", "degenerate NLR")) %>% # Select only NLRs
mutate(ID = case_when(str_detect(Domain_simplified, "O") ~ "YES", # Add a column for whether the NLR contains a putative integrated domain
TRUE ~ "NO")) %>%
group_by(RefSeq, Locus) %>%
arrange(desc(ID),
desc(Status)) %>%
slice(1) %>% # Select a single splice-variant per locus
ungroup %>%
filter(!grepl("SUPPRESSED", Species)) %>% # Remove the suppressed RefSeq genomes
group_by(RefSeq, Species, ID) %>%
summarise(count = n()) %>%
mutate(perc = (count/sum(count))*100)
head(RefSeq_NLR_LOCUS)
#> # A tibble: 6 × 5
#> # Groups: RefSeq, Species [3]
#> RefSeq Species ID count perc
#> <chr> <chr> <chr> <int> <dbl>
#> 1 GCF_000001735.4 Arabidopsis thaliana NO 167 95.4
#> 2 GCF_000001735.4 Arabidopsis thaliana YES 8 4.57
#> 3 GCF_000002425.4 Physcomitrella patens NO 39 68.4
#> 4 GCF_000002425.4 Physcomitrella patens YES 18 31.6
#> 5 GCF_000002775.4 Populus trichocarpa NO 454 94.2
#> 6 GCF_000002775.4 Populus trichocarpa YES 28 5.81
ggplot(data = RefSeq_NLR_LOCUS, aes(x = Species, y = count, fill = ID)) +
geom_bar(stat = "identity") +
scale_x_discrete(limits = c("Physcomitrella patens", "Selaginella moellendorffii", # non-flowering plants
"Amborella trichopoda", "Nymphaea colorata", # Magnoliopsida
"Dioscorea rotundata", "Asparagus officinalis", "Dendrobium catenatum", "Phalaenopsis equestris", # Liliopsida, Petrosaviidae
"Elaeis guineensis", "Phoenix dactylifera", # Liliopsida, Petrosaviidae, Arecales
"Musa acuminata", # Liliopsida, Petrosaviidae, Zingiberales
"Ananas comosus", # Liliopsida, Petrosaviidae, Poales, Bromeliaceae
"Panicum hallii", "Panicum virgatum", "Setaria italica", "Setaria viridis", "Sorghum bicolor", "Zea mays", # Liliopsida, Petrosaviidae, Poales, Poaceae, PACMAD clade
"Oryza brachyantha", "Oryza sativa", "Brachypodium distachyon", "Aegilops tauschii", "Triticum dicoccoides", # Liliopsida, Petrosaviidae, Poales, Poaceae, BOP clade
"Nelumbo nucifera", "Papaver somniferum", # Eudicotyledons
"Beta vulgaris", "Chenopodium quinoa", "Spinacia oleracea", # Eudicotyledons, Superasterids, Caryophyllales
"Camellia sinensis", # Eudicotyledons, Superasterids, Asterids, Ericales
"Daucus carota", "Cynara cardunculus", "Helianthus annuus", "Lactuca sativa", # Eudicotyledons, Superasterids, Asterids, campanulids
"Coffea eugenioides", "Coffea arabica", # Eudicotyledons, Superasterids, Asterids, Lamiids, Gentianales
"Olea europaea", "Sesamum indicum", "Erythranthe guttata", # Eudicotyledons, Superasterids, Asterids, Lamiids, Lamiales
"Ipomoea nil", "Ipomoea triloba", # Eudicotyledons, Superasterids, Asterids, Lamiids, Solanales, Convolvulaceae
"Nicotiana tomentosiformis", "Nicotiana sylvestris", "Nicotiana tabacum", "Nicotiana attenuata", "Capsicum annuum", "Solanum tuberosum", "Solanum pennellii", "Solanum lycopersicum", # Eudicotyledons, Superasterids, Asterids, Lamiids, Solanales, Solanaceae
"Vitis riparia", "Vitis vinifera", # Eudicotyledons, Rosids, rosids incertae sedis, Vitales
"Tripterygium wilfordii", # Eudicotyledons, Rosids, Fabids, Celastrales
"Hevea brasiliensis", "Jatropha curcas", "Manihot esculenta", "Ricinus communis", "Populus trichocarpa", "Populus euphratica", "Populus alba", # Eudicotyledons, Rosids, Fabids, Malpighiales
"Prosopis alba", "Lupinus angustifolius", "Arachis hypogaea", "Arachis duranensis", "Arachis ipaensis", "Cicer arietinum", "Medicago truncatula", "Abrus precatorius", "Vigna radiata", "Vigna angularis", "Vigna unguiculata", "Cajanus cajan", "Phaseolus vulgaris", "Glycine soja", "Glycine max", # Eudicotyledons, Rosids, Fabids, Fabales
"Fragaria vesca", "Rosa chinensis", "Pyrus x bretschneideri", "Malus domestica", "Prunus dulcis", "Prunus avium", "Prunus mume", "Prunus persica", "Ziziphus jujuba", "Cannabis sativa", "Morus notabilis", # Eudicotyledons, Rosids, Fabids, Rosales
"Juglans regia", "Juglans microcarpa x Juglans regia", "Quercus lobata", "Quercus suber", # Eudicotyledons, Rosids, Fabids, Fagales
"Momordica charantia", "Cucurbita pepo", "Cucurbita moschata", "Cucurbita maxima", "Benincasa hispida", "Cucumis melo", "Cucumis sativus", # Eudicotyledons, Rosids, Fabids, Cucurbitales
"Punica granatum", "Rhodamnia argentea", "Syzygium oleosum", "Eucalyptus grandis", # Eudicotyledons, Rosids, Malvids, Myrtales
"Pistacia vera", "Citrus sinensis", "Citrus clementina", # Eudicotyledons, Rosids, Malvids, Sapindales
"Durio zibethinus", "Herrania umbratica", "Theobroma cacao", "Hibiscus syriacus", "Gossypium arboreum", "Gossypium raimondii", "Gossypium hirsutum", # Eudicotyledons, Rosids, Malvids, Malvales
"Tarenaya hassleriana", "Carica papaya", "Eutrema salsugineum", "Raphanus sativus", "Brassica rapa", "Brassica napus", "Brassica oleracea", "Capsella rubella", "Camelina sativa", "Arabidopsis lyrata", "Arabidopsis thaliana" # Eudicotyledons, Rosids, Malvids, Brassicales
)) +
scale_y_continuous(breaks = c(0, 250, 500, 750, 1000, 1250)) +
scale_fill_manual(values = c("YES" = "#F0027F", "NO" = "#386CB0")) +
coord_flip() +
theme(panel.background = element_rect(fill = NA),
panel.grid.major = element_line(colour = "#BFBFBF", size = 0.6),
panel.grid.major.y = element_line(colour = NA),
axis.line.x = element_line(color = "#BFBFBF", size = 0.6),
axis.ticks.x = element_line(color = "#BFBFBF", size = 0.6),
axis.ticks.y = element_line(color = "#BFBFBF", size = 0.6),
axis.text.y = element_text(angle = 0, hjust = 1, size = 8),
axis.title.y=element_blank())
We can also plot the types of integrated domains found
RefSeq_NLR_ID_DEDUP <- RefSeq_NLR %>%
filter(!grepl("SUPPRESSED", Species), # Remove the suppressed RefSeq genomes
Status %in% c("NLR", "degenerate NLR"), # Filter for NLRs
grepl("O", Domain_simplified), # Extract NLRs containing a potential integrated domain
!type %in% c("CHAIN", "MOTIF", "REGION")) %>% # Remove CHAIN and MOTIF and REGION (canonical NLR domains)
separate_rows(Domain, sep = "\\)\\(") %>% # Separate out the domains
mutate(Domain = str_replace(Domain, "\\(", ""),
Domain = str_replace(Domain, "\\)", "")) %>%
filter(!Domain %in% c("R1", "RPW8", "TIR", "CC", "NBARC", "PLOOP", "LRR", "CJID")) %>% # Remove the canonical NLR domains from the domain architecture analysis
group_by(RefSeq, Locus) %>%
distinct(Domain, .keep_all = TRUE) %>% # Deduplicate the integrated domains per locus.
group_by(RefSeq, Species, Domain) %>%
summarise(count = n()) %>%
mutate(perc = (count/sum(count))*100,
Domain = factor(Domain, levels = c("WRKY", "TRX", "PLOOP-OTHER", "PKin", "HMA", "Exo70", "BED", "a/b", "JAC", "BRX", "WD40", "TPR", "ARM", "ANK", "OTHER", "TRANSPOSON")))
head(RefSeq_NLR_ID_DEDUP)
#> # A tibble: 6 × 5
#> # Groups: RefSeq, Species [2]
#> RefSeq Species Domain count perc
#> <chr> <chr> <fct> <int> <dbl>
#> 1 GCF_000001735.4 Arabidopsis thaliana BRX 1 10
#> 2 GCF_000001735.4 Arabidopsis thaliana OTHER 4 40
#> 3 GCF_000001735.4 Arabidopsis thaliana PKin 1 10
#> 4 GCF_000001735.4 Arabidopsis thaliana PLOOP-OTHER 1 10
#> 5 GCF_000001735.4 Arabidopsis thaliana WRKY 3 30
#> 6 GCF_000002425.4 Physcomitrella patens OTHER 1 5.56
ggplot(data = RefSeq_NLR_ID_DEDUP, aes(x = Species, y = count, fill = Domain)) +
geom_bar(stat = "identity") +
scale_x_discrete(limits = c("Physcomitrella patens", "Selaginella moellendorffii", # non-flowering plants
"Amborella trichopoda", "Nymphaea colorata", # Magnoliopsida
"Dioscorea rotundata", "Asparagus officinalis", "Dendrobium catenatum", "Phalaenopsis equestris", # Liliopsida, Petrosaviidae
"Elaeis guineensis", "Phoenix dactylifera", # Liliopsida, Petrosaviidae, Arecales
"Musa acuminata", # Liliopsida, Petrosaviidae, Zingiberales
"Ananas comosus", # Liliopsida, Petrosaviidae, Poales, Bromeliaceae
"Panicum hallii", "Panicum virgatum", "Setaria italica", "Setaria viridis", "Sorghum bicolor", "Zea mays", # Liliopsida, Petrosaviidae, Poales, Poaceae, PACMAD clade
"Oryza brachyantha", "Oryza sativa", "Brachypodium distachyon", "Aegilops tauschii", "Triticum dicoccoides", # Liliopsida, Petrosaviidae, Poales, Poaceae, BOP clade
"Nelumbo nucifera", "Papaver somniferum", # Eudicotyledons
"Beta vulgaris", "Chenopodium quinoa", "Spinacia oleracea", # Eudicotyledons, Superasterids, Caryophyllales
"Camellia sinensis", # Eudicotyledons, Superasterids, Asterids, Ericales
"Daucus carota", "Cynara cardunculus", "Helianthus annuus", "Lactuca sativa", # Eudicotyledons, Superasterids, Asterids, campanulids
"Coffea eugenioides", "Coffea arabica", # Eudicotyledons, Superasterids, Asterids, Lamiids, Gentianales
"Olea europaea", "Sesamum indicum", "Erythranthe guttata", # Eudicotyledons, Superasterids, Asterids, Lamiids, Lamiales
"Ipomoea nil", "Ipomoea triloba", # Eudicotyledons, Superasterids, Asterids, Lamiids, Solanales, Convolvulaceae
"Nicotiana tomentosiformis", "Nicotiana sylvestris", "Nicotiana tabacum", "Nicotiana attenuata", "Capsicum annuum", "Solanum tuberosum", "Solanum pennellii", "Solanum lycopersicum", # Eudicotyledons, Superasterids, Asterids, Lamiids, Solanales, Solanaceae
"Vitis riparia", "Vitis vinifera", # Eudicotyledons, Rosids, rosids incertae sedis, Vitales
"Tripterygium wilfordii", # Eudicotyledons, Rosids, Fabids, Celastrales
"Hevea brasiliensis", "Jatropha curcas", "Manihot esculenta", "Ricinus communis", "Populus trichocarpa", "Populus euphratica", "Populus alba", # Eudicotyledons, Rosids, Fabids, Malpighiales
"Prosopis alba", "Lupinus angustifolius", "Arachis hypogaea", "Arachis duranensis", "Arachis ipaensis", "Cicer arietinum", "Medicago truncatula", "Abrus precatorius", "Vigna radiata", "Vigna angularis", "Vigna unguiculata", "Cajanus cajan", "Phaseolus vulgaris", "Glycine soja", "Glycine max", # Eudicotyledons, Rosids, Fabids, Fabales
"Fragaria vesca", "Rosa chinensis", "Pyrus x bretschneideri", "Malus domestica", "Prunus dulcis", "Prunus avium", "Prunus mume", "Prunus persica", "Ziziphus jujuba", "Cannabis sativa", "Morus notabilis", # Eudicotyledons, Rosids, Fabids, Rosales
"Juglans regia", "Juglans microcarpa x Juglans regia", "Quercus lobata", "Quercus suber", # Eudicotyledons, Rosids, Fabids, Fagales
"Momordica charantia", "Cucurbita pepo", "Cucurbita moschata", "Cucurbita maxima", "Benincasa hispida", "Cucumis melo", "Cucumis sativus", # Eudicotyledons, Rosids, Fabids, Cucurbitales
"Punica granatum", "Rhodamnia argentea", "Syzygium oleosum", "Eucalyptus grandis", # Eudicotyledons, Rosids, Malvids, Myrtales
"Pistacia vera", "Citrus sinensis", "Citrus clementina", # Eudicotyledons, Rosids, Malvids, Sapindales
"Durio zibethinus", "Herrania umbratica", "Theobroma cacao", "Hibiscus syriacus", "Gossypium arboreum", "Gossypium raimondii", "Gossypium hirsutum", # Eudicotyledons, Rosids, Malvids, Malvales
"Tarenaya hassleriana", "Carica papaya", "Eutrema salsugineum", "Raphanus sativus", "Brassica rapa", "Brassica napus", "Brassica oleracea", "Capsella rubella", "Camelina sativa", "Arabidopsis lyrata", "Arabidopsis thaliana" # Eudicotyledons, Rosids, Malvids, Brassicales
)) +
scale_fill_manual(values = colorRampPalette(RColorBrewer::brewer.pal(12, "Accent"))(length(unique(RefSeq_NLR_ID_DEDUP$Domain)))) +
scale_y_continuous(breaks = c(0, 25, 50, 75, 100)) +
coord_flip() +
theme(panel.background = element_rect(fill = NA),
panel.grid.major = element_line(colour = "#BFBFBF", size = 0.6),
panel.grid.major.y = element_line(colour = NA),
axis.line.x = element_line(color = "#BFBFBF", size = 0.6),
axis.ticks.x = element_line(color = "#BFBFBF", size = 0.6),
axis.ticks.y = element_line(color = "#BFBFBF", size = 0.6),
axis.text.y = element_text(angle = 0, hjust = 1, size = 8),
axis.title.y=element_blank())
#> Warning in RColorBrewer::brewer.pal(12, "Accent"): n too large, allowed maximum for palette Accent is 8
#> Returning the palette you asked for with that many colors