Last updated: 2020-06-08
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Knit directory: Comparative_APA/analysis/
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Rmd | 085794d | brimittleman | 2020-06-08 | add trpseq analysis |
library(tidyverse)
── Attaching packages ──────────────── tidyverse 1.2.1 ──
✔ ggplot2 3.1.1 ✔ purrr 0.3.2
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── Conflicts ─────────────────── tidyverse_conflicts() ──
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library(ggpubr)
Loading required package: magrittr
Attaching package: 'magrittr'
The following object is masked from 'package:purrr':
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The following object is masked from 'package:tidyr':
extract
THe trip seq paper has a method to explore 3’ UTR characteristics from a bed file
https://github.com/stephenfloor/tripseq-analysis Test transcriptome_properties.py
I need the name of the genome and a bed file. I can start by testing on the ortho 3’ UTRs
clone code/tripseq-analysis python 2
source ~/activate_anaconda_python2.sh
module load bedtools
#in trip
python transcriptome_properties.py -i ../../data/orthoUTR/HumanDistal3UTR.sort.bed -g /project2/gilad/kenneth/References/human/genome/hg38.fa --au-elements
This gets the au rich element %.
can do this with human and chimp:
python transcriptome_properties.py -i ../../data/orthoUTR/HumanDistal3UTR.sort.bed -g /project2/gilad/kenneth/References/human/genome/hg38.fa --au-elements -o ../../data/orthoUTR/HumanOrthoUTR_AUrich
python transcriptome_properties.py -i ../../data/orthoUTR/ChimpDistal3UTR.sort.bed -g /project2/gilad/briana/genome_anotation_data/Chimp_genome/panTro6.fa --au-elements -o ../../data/orthoUTR/ChimpOrthoUTR_AUrich
HumanOrthUTR_au=read.csv("../data/orthoUTR/HumanOrthoUTR_AUrich_au_elements.csv",header = T, stringsAsFactors = F)
ChimpOrthoUTR_au=read.csv("../data/orthoUTR/ChimpOrthoUTR_AUrich_au_elements.csv",header = T, stringsAsFactors = F)
Corr:
HumanOrthUTR_au_sm=HumanOrthUTR_au %>% select(transcriptID, au_element_frac) %>% rename(HumanAU=au_element_frac)
ChimpOrthUTR_au_sm=ChimpOrthoUTR_au %>% select(transcriptID, au_element_frac) %>% rename(ChimpAU=au_element_frac)
BothAU=HumanOrthUTR_au_sm %>% inner_join(ChimpOrthUTR_au_sm, by="transcriptID")
cor.test(BothAU$ChimpAU, BothAU$HumanAU)
Pearson's product-moment correlation
data: BothAU$ChimpAU and BothAU$HumanAU
t = 584.48, df = 15739, p-value < 2.2e-16
alternative hypothesis: true correlation is not equal to 0
95 percent confidence interval:
0.9770321 0.9784085
sample estimates:
cor
0.9777308
AU number:
HumanOrthUTR_au_Numsm=HumanOrthUTR_au %>% select(transcriptID, au_element_count) %>% rename(HumanAU=au_element_count)
ChimpOrthUTR_au_Numsm=ChimpOrthoUTR_au %>% select(transcriptID, au_element_count) %>% rename(ChimpAU=au_element_count)
BothAUNum=HumanOrthUTR_au_Numsm %>% inner_join(ChimpOrthUTR_au_Numsm, by="transcriptID")
cor.test(BothAU$ChimpAU, BothAU$HumanAU)
Pearson's product-moment correlation
data: BothAU$ChimpAU and BothAU$HumanAU
t = 584.48, df = 15739, p-value < 2.2e-16
alternative hypothesis: true correlation is not equal to 0
95 percent confidence interval:
0.9770321 0.9784085
sample estimates:
cor
0.9777308
same high correlation…
ggplot(BothAU, aes(x=HumanAU, y=ChimpAU)) + geom_point() + geom_abline(slope=1) + geom_density_2d()
Color by dAPA and diff iso diversity.
Meta=read.table("../data/PAS_doubleFilter/PAS_5perc_either_HumanCoord_BothUsage_meta_doubleFilter.txt", header = T, stringsAsFactors = F)
Meta_genes= Meta %>% select(gene) %>% unique()
Meta_PAS=Meta %>% select(PAS,gene)
dAPAGenes=read.table("../data/DiffIso_Nuclear_DF/SignifianceEitherGENES_Nuclear.txt", header = T, stringsAsFactors = F)
dAPAPAS=read.table("../data/DiffIso_Nuclear_DF/AllPAS_withGeneSig.txt", header = T, stringsAsFactors = F) %>% inner_join(Meta, by=c("chr","start", "end","gene")) %>% select(PAS,gene,SigPAU2 )
dAPAPAS_genes= dAPAPAS %>% select(gene) %>% unique()
dAPATestedGenes= dAPAPAS %>% select(gene) %>% unique() %>% mutate(dAPA=ifelse(gene %in% dAPAGenes$gene,"Yes", "No"))
dICdata= read.table("../data/IndInfoContent/SimpsonMedianSignificance.txt", header = T, stringsAsFactors = F)%>% select(sIC,gene)
dICdata_sig= dICdata %>% filter(sIC=="Yes")
dAPAandDic= dICdata %>% inner_join(dAPATestedGenes,by="gene") %>% mutate(Both=ifelse(sIC=="Yes" & dAPA=="Yes", "Yes","No"),OnlyIC=ifelse(sIC=="Yes" & dAPA=="No", "Yes","No"),OnlyAPA=ifelse(sIC=="No" & dAPA=="Yes", "Yes","No"))
dAPAonly=dAPAandDic %>% filter(dAPA=="Yes") %>% select(gene) %>% mutate(set="dAPA")
both=dAPAandDic %>% filter(Both=="Yes") %>% select(gene)%>% mutate(set="Both")
IDonly=dAPAandDic %>% filter(OnlyIC=="Yes") %>% select(gene)%>% mutate(set="ID")
Allset=dAPAonly %>% bind_rows(both) %>% bind_rows(IDonly)
Nonegenes= Meta_PAS %>% select(gene) %>% unique()%>% anti_join(Allset, by="gene") %>% mutate(set="None")
#no diff:
Allset_andnon=Allset %>% bind_rows(Nonegenes)
BothAU_fix= BothAU %>% separate(transcriptID, into=c("gene", "strand"), sep="\\(") %>% inner_join(Allset_andnon, by="gene") %>% mutate(anyDiff=ifelse(set=="None", "No", "Yes"))
BothAU_fix %>% group_by(set) %>% summarise(n())
# A tibble: 4 x 2
set `n()`
<chr> <int>
1 Both 329
2 dAPA 1281
3 ID 345
4 None 5835
Do dAPA genes have higher AU
BothAU_fix_g= BothAU_fix %>% gather("Species", "AU", -gene, -strand, -set,-anyDiff)
ggplot(BothAU_fix_g, aes(x=AU,col=set)) +stat_ecdf() + facet_grid(~Species)
Version | Author | Date |
---|---|---|
4ff1c53 | brimittleman | 2020-06-08 |
box plots:
ggplot(BothAU_fix_g, aes(y=AU,x=set,fill=set)) +geom_boxplot() + facet_grid(~Species) + stat_compare_means()
ggplot(BothAU_fix_g, aes(y=AU,x=anyDiff,fill=anyDiff)) +geom_boxplot() + facet_grid(~Species) + stat_compare_means(method.args = list(alternative = "greater"))
BothAU_fix_g %>% group_by(Species,anyDiff) %>% summarise(mean(AU))
# A tibble: 4 x 3
# Groups: Species [2]
Species anyDiff `mean(AU)`
<chr> <chr> <dbl>
1 ChimpAU No 0.0151
2 ChimpAU Yes 0.0162
3 HumanAU No 0.0152
4 HumanAU Yes 0.0163
BothAU_fix_g %>% group_by(Species,set) %>% summarise(mean(AU))
# A tibble: 8 x 3
# Groups: Species [2]
Species set `mean(AU)`
<chr> <chr> <dbl>
1 ChimpAU Both 0.0156
2 ChimpAU dAPA 0.0166
3 ChimpAU ID 0.0157
4 ChimpAU None 0.0151
5 HumanAU Both 0.0158
6 HumanAU dAPA 0.0166
7 HumanAU ID 0.0155
8 HumanAU None 0.0152
dAPA are the higher genes:
genes with expression diff:
nameID=read.table("../../genome_anotation_data/ensemble_to_genename.txt",sep="\t", header = T, stringsAsFactors = F) %>% dplyr::select(Gene_stable_ID, Gene.name)
DiffExp=read.table("../data/DiffExpression/DEtested_allres.txt",stringsAsFactors = F,header = F, col.names = c("Gene_stable_ID" ,"logFC" ,"AveExpr" , "t" , "P.Value" , "adj.P.Val", "B" )) %>% inner_join(nameID,by="Gene_stable_ID") %>% dplyr::rename('gene'=Gene.name) %>% dplyr::select(-Gene_stable_ID) %>% mutate(DE=ifelse(adj.P.Val<.05, "Yes", "No")) %>% select(gene,DE)
BothAU_fix_g_DE= BothAU_fix_g %>% inner_join(DiffExp,by="gene")
ggplot(BothAU_fix_g_DE, aes(y=AU,x=DE,fill=DE)) +geom_boxplot() + facet_grid(anyDiff~Species) + stat_compare_means()
Version | Author | Date |
---|---|---|
4ff1c53 | brimittleman | 2020-06-08 |
genes with differential translation:
Ribo=read.table("../data/Wang_ribo/Additionaltable5_translationComparisons.txt",header = T, stringsAsFactors = F) %>% rename("Gene_stable_ID"= ENSG) %>% inner_join(nameID,by="Gene_stable_ID") %>% dplyr::select(Gene.name, HvC.beta, HvC.pvalue, HvC.FDR) %>% rename("gene"=Gene.name) %>% mutate(dTE=ifelse(HvC.FDR <0.05, "Yes","No"))
RiboSmall= Ribo %>% select(gene,dTE)
BothAU_fix_g_TE= BothAU_fix_g %>% inner_join(RiboSmall,by="gene")
ggplot(BothAU_fix_g_TE, aes(y=AU,x=dTE,fill=dTE)) +geom_boxplot() + facet_grid(anyDiff~Species) + stat_compare_means()
Version | Author | Date |
---|---|---|
4ff1c53 | brimittleman | 2020-06-08 |
AU for the ortho UTRs are not different based on DE or translation status.
filter to dAPA genes and see if they are different in expression and translation:
BothAU_fix_g_DE_dAPA= BothAU_fix_g_DE %>% filter(set=="dAPA")
ggplot(BothAU_fix_g_DE_dAPA, aes(y=AU,x=DE,fill=DE)) +geom_boxplot() + facet_grid(~Species) + stat_compare_means()
BothAU_fix_g_TE_dAPA= BothAU_fix_g_TE %>% filter(set=="dAPA")
ggplot(BothAU_fix_g_TE_dAPA, aes(y=AU,x=dTE,fill=dTE)) +geom_boxplot() + facet_grid(~Species) + stat_compare_means()
ok so only significant relationship here is higher AU proportion in ortho 3’ UTRs for dAPA genes.
difference in dominance:
HumanRes=read.table("../data/DomDefGreaterX/Human_AllGenes_DiffTop.txt", col.names = c("Human_PAS", "gene","Human_DiffDom"),stringsAsFactors = F)
ChimpRes=read.table("../data/DomDefGreaterX/Chimp_AllGenes_DiffTop.txt", col.names = c("Chimp_PAS", "gene","Chimp_DiffDom"),stringsAsFactors = F)
BothRes=HumanRes %>% inner_join(ChimpRes,by="gene")
BothRes_10=BothRes %>% filter(Chimp_DiffDom >=0.1 | Human_DiffDom>=0.1) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=10)
BothRes_20=BothRes %>% filter(Chimp_DiffDom >=0.2 | Human_DiffDom>=0.2) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=20)
BothRes_30=BothRes %>% filter(Chimp_DiffDom >=0.3 | Human_DiffDom>=0.3) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=30)
BothRes_40=BothRes %>% filter(Chimp_DiffDom >=0.4 | Human_DiffDom>=0.4) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=40)
BothRes_50=BothRes %>% filter(Chimp_DiffDom >=0.5 | Human_DiffDom>=0.5) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=50)
BothRes_60=BothRes %>% filter(Chimp_DiffDom >=0.6 | Human_DiffDom>=0.6) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=60)
BothRes_70=BothRes %>% filter(Chimp_DiffDom >=0.7 | Human_DiffDom>=0.7) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=70)
BothRes_80=BothRes %>% filter(Chimp_DiffDom >=0.8 | Human_DiffDom>=0.8) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=80)
BothRes_90=BothRes %>% filter(Chimp_DiffDom >=0.9 | Human_DiffDom>=0.9) %>% mutate(Set= ifelse(Human_PAS==Chimp_PAS,"Same", "Different"),cut=90)
BothResAll=BothRes_10 %>% bind_rows(BothRes_20) %>% bind_rows(BothRes_30) %>% bind_rows(BothRes_40) %>% bind_rows(BothRes_50) %>% bind_rows(BothRes_60) %>% bind_rows(BothRes_70) %>% bind_rows(BothRes_80) %>% bind_rows(BothRes_90)
plotting au in human for each:
BothResAll_au= BothResAll %>% inner_join(BothAU_fix,by="gene")
BothResAll_au$cut=as.factor(BothResAll_au$cut)
ggplot(BothResAll_au,aes(x=cut, by=Set, y=HumanAU,fill=Set)) + geom_boxplot() + stat_compare_means(aes(label = ..p.signif..))
DP not DE:
ProtInfo=read.table("../data/PTM/ProtLength.txt", sep = "\t",stringsAsFactors = F,header = T,col.names = c("entry","organism", "nAA", "gene")) %>% select(nAA, gene)
Prot= read.table("../data/Khan_prot/ProtData_effectSize.txt",header = T,stringsAsFactors = F) %>% mutate(dP=ifelse(pval<0.05, "Yes", "No")) %>% filter(dP=="Yes")
Interactions=read.table("../data/bioGRID/GeneswInteractions.txt",stringsAsFactors = F, header = T) %>% inner_join(ProtInfo, by="gene")%>% mutate(NormInter=nInt/nAA)
#DiffExp$gene, DT=Ribo$gene, DP=Prot$gene
dAPAandDic_wP=dAPAandDic %>% mutate(dE=ifelse(gene %in%DiffExp$gene, "Yes", "No" ), dP=ifelse(gene %in%Prot$gene,"Yes","No" ), dPnotDE=ifelse(dE=="No"&dP=="Yes", "Yes","No")) %>% inner_join(Interactions, by="gene") %>% inner_join(BothAU_fix,by="gene")
ggplot(dAPAandDic_wP, aes(x=dPnotDE, y=HumanAU)) + geom_boxplot()
only one gene with the pattern and an ortho 3’ UTR.
Human 3’ UTRs:
Merge from refseq anno:
mkdir ../data/HumanUTR
bedtools merge -s -i ../data/orthoUTR/g38_ncbiRefseq_Formatted_Allannotation_UTR3.bed -c 4,5,6 -o distinct,mean,distinct > ../data/HumanUTR/Human3UTR.merged.bed
still multiple per genes. take most 3’ ?
humanMergeutr= read.table("../data/HumanUTR/Human3UTR.merged.bed", col.names = c('chr','start','end','gene','score','strand'), stringsAsFactors = F) %>% group_by(gene)
humanMergeutrpos= humanMergeutr %>% filter(strand=="+") %>% group_by(gene) %>% top_n(1,start)
humanMergeutrneg= humanMergeutr %>% filter(strand=="-") %>% group_by(gene) %>% top_n(-1,start)
humanDistalboth=humanMergeutrpos %>% bind_rows(humanMergeutrneg)
write.table(humanDistalboth, "../data/HumanUTR/HumanDistal3UTR.bed", col.names = F, row.names = F, quote = F, sep="\t")
Run these through AU:
python transcriptome_properties.py -i../../data/HumanUTR/HumanDistal3UTR.bed -g /project2/gilad/kenneth/References/human/genome/hg38.fa --au-elements -o ../../data/HumanUTR/HumanAllUTR_AUrich
HumanALLUTR_au=read.csv("../data/HumanUTR/HumanAllUTR_AUrich_au_elements.csv",header = T, stringsAsFactors = F) %>% separate(transcriptID, into=c("geneloc", "strand"), sep="\\(") %>% separate(geneloc, into=c("loc","gene"),sep=":") %>% na.omit()
Warning: Expected 2 pieces. Missing pieces filled with `NA` in 611 rows
[63, 64, 65, 66, 135, 136, 409, 410, 463, 464, 611, 612, 691, 692, 974,
975, 1074, 1075, 1466, 1467, ...].
Warning: Expected 2 pieces. Missing pieces filled with `NA` in 302 rows
[64, 66, 136, 410, 464, 612, 692, 975, 1075, 1467, 1747, 1955, 2461, 2566,
2620, 2688, 2801, 2970, 3002, 3029, ...].
missing pieces were overlapping.
dAPAandDic_wPAU=dAPAandDic %>% mutate(dE=ifelse(gene %in%DiffExp$gene, "Yes", "No" ), dP=ifelse(gene %in%Prot$gene,"Yes","No" ), dPnotDE=ifelse(dE=="No"&dP=="Yes", "Yes","No")) %>% inner_join(Interactions, by="gene") %>% inner_join(HumanALLUTR_au, by="gene")
dAPAandDic_wPAU$au_element_frac= as.numeric(as.character(dAPAandDic_wPAU$au_element_frac))
ggplot(dAPAandDic_wPAU, aes(x=dPnotDE, y=au_element_frac)) + geom_boxplot()
dAPAandDic_wPAU %>% group_by(dPnotDE) %>% summarise(n())
# A tibble: 2 x 2
dPnotDE `n()`
<chr> <int>
1 No 7132
2 Yes 2
only 2…
Allset_andnon_au= Allset_andnon %>% inner_join(HumanALLUTR_au, by="gene")
Allset_andnon_au$au_element_frac= as.numeric(as.character(Allset_andnon_au$au_element_frac))
ggplot(Allset_andnon_au, aes(x=set,fill=set,y=au_element_frac)) + geom_boxplot() + stat_compare_means()
Allset_andnon_au %>% group_by(set ) %>% summarise(mean(au_element_frac))
# A tibble: 4 x 2
set `mean(au_element_frac)`
<chr> <dbl>
1 Both 0.0150
2 dAPA 0.0161
3 ID 0.0162
4 None 0.0154
Allset_andnon_aude=Allset_andnon_au %>% inner_join(DiffExp,by="gene")
ggplot(Allset_andnon_aude, aes(x=set,fill=set,y=au_element_frac,by=DE)) + geom_boxplot() + stat_compare_means()
Allset_andnon_auTE=Allset_andnon_au %>% inner_join(RiboSmall,by="gene")
ggplot(Allset_andnon_auTE, aes(x=set,fill=set,y=au_element_frac,by=dTE)) + geom_boxplot() + stat_compare_means()
ok there are no interesting relationships with this… I would have to proportion the UTRs probably…
number:
Allset_andnon_au$au_element_count=as.numeric(Allset_andnon_au$au_element_count)
ggplot(Allset_andnon_au, aes(x=set,fill=set,y=au_element_count)) + geom_boxplot() + stat_compare_means() +ylim(c(0,100))
Warning: Removed 1 rows containing non-finite values (stat_boxplot).
Warning: Removed 1 rows containing non-finite values (stat_compare_means).
Allset_andnon_auany= Allset_andnon_au %>% mutate(AnyAPA=ifelse(set=="None","No", "Yes"))
ggplot(Allset_andnon_auany, aes(x=AnyAPA,fill=AnyAPA,y=au_element_count)) + geom_boxplot() + stat_compare_means() +ylim(c(0,100))
Warning: Removed 1 rows containing non-finite values (stat_boxplot).
Warning: Removed 1 rows containing non-finite values (stat_compare_means).
ggplot(Allset_andnon_auany, aes(x=AnyAPA,fill=AnyAPA,y=au_element_frac)) + geom_boxplot() + stat_compare_means(method.args = list(alternative = "less"))
Allset_andnon_auany_de= Allset_andnon_auany %>% inner_join(DiffExp,by="gene")
ggplot(Allset_andnon_auany_de, aes(x=DE,fill=AnyAPA,y=au_element_count)) + geom_boxplot() + stat_compare_means() +ylim(c(0,100))
dAPA have more AU elements but the other relationships dont make much sense. nothing is strong.. on hold for now.
sessionInfo()
R version 3.5.1 (2018-07-02)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Scientific Linux 7.4 (Nitrogen)
Matrix products: default
BLAS/LAPACK: /software/openblas-0.2.19-el7-x86_64/lib/libopenblas_haswellp-r0.2.19.so
locale:
[1] LC_CTYPE=en_US.UTF-8 LC_NUMERIC=C
[3] LC_TIME=en_US.UTF-8 LC_COLLATE=en_US.UTF-8
[5] LC_MONETARY=en_US.UTF-8 LC_MESSAGES=en_US.UTF-8
[7] LC_PAPER=en_US.UTF-8 LC_NAME=C
[9] LC_ADDRESS=C LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
attached base packages:
[1] stats graphics grDevices utils datasets methods base
other attached packages:
[1] ggpubr_0.2 magrittr_1.5 forcats_0.3.0 stringr_1.3.1
[5] dplyr_0.8.0.1 purrr_0.3.2 readr_1.3.1 tidyr_0.8.3
[9] tibble_2.1.1 ggplot2_3.1.1 tidyverse_1.2.1
loaded via a namespace (and not attached):
[1] tidyselect_0.2.5 reshape2_1.4.3 haven_1.1.2 lattice_0.20-38
[5] colorspace_1.3-2 generics_0.0.2 htmltools_0.3.6 yaml_2.2.0
[9] utf8_1.1.4 rlang_0.4.0 later_0.7.5 pillar_1.3.1
[13] glue_1.3.0 withr_2.1.2 modelr_0.1.2 readxl_1.1.0
[17] plyr_1.8.4 munsell_0.5.0 gtable_0.2.0 workflowr_1.6.0
[21] cellranger_1.1.0 rvest_0.3.2 evaluate_0.12 labeling_0.3
[25] knitr_1.20 httpuv_1.4.5 fansi_0.4.0 broom_0.5.1
[29] Rcpp_1.0.4.6 promises_1.0.1 scales_1.0.0 backports_1.1.2
[33] jsonlite_1.6 fs_1.3.1 hms_0.4.2 digest_0.6.18
[37] stringi_1.2.4 grid_3.5.1 rprojroot_1.3-2 cli_1.1.0
[41] tools_3.5.1 lazyeval_0.2.1 crayon_1.3.4 whisker_0.3-2
[45] pkgconfig_2.0.2 MASS_7.3-51.1 xml2_1.2.0 lubridate_1.7.4
[49] assertthat_0.2.0 rmarkdown_1.10 httr_1.3.1 rstudioapi_0.10
[53] R6_2.3.0 nlme_3.1-137 git2r_0.26.1 compiler_3.5.1