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dts-seq-plot.r
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dts-seq-plot.r
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# Script for Termination Signal (ts) calculation and plots
# Daniel Gautheret, 2018
rm(list=ls())
library (Biostrings)
# ---------- INPUT FILES ---------------
datadir="./"
covdir="5_coverage/"
# tRNA coordinates and names
trnacoordF<-paste0(datadir,"6_tRNA_modification/NC_000913_tRNA.tsv")
# coli fasta sequence imported from:
# ftp://ftp.ncbi.nlm.nih.gov/genomes/all/GCF/000/005/845/GCF_000005845.2_ASM584v2/GCF_000005845.2_ASM584v2_genomic.fna.gz
# (unzipped)
genomeF<-paste0(datadir, "1_rawData/GCF_000005845.2_ASM584v2_genomic.fna")
# Modomics modified bases (Fasta-like but shows only mod bases)
modomicsF<-paste0(datadir, "6_tRNA_modification/bmModomics_with_tmRNA.fasta")
# list of tRNAs for printing and names of "lead" tRNAs
trnaF=paste0(datadir,"7_termination_signal/trnaprint.txt")
# Genome mapping coverage files (BigWig-like but w/ 2 strands)
# A 2-column file with one line per genome position
# column 1: coverage for + strand
# column 2: coverage for minus strand
covF=c(
"A4-NT_CCATGG_depth_fr.txt",
"A3-D_CCATGG_depth_fr.txt",
"A5-nD_CCATGG_depth_fr.txt",
"B4-NT_CCATGG_depth_fr.txt",
"B3-D_CCATGG_depth_fr.txt",
"B5-nD_CCATGG_depth_fr.txt",
"C4-NT_CCATGG_depth_fr.txt",
"C3-D_CCATGG_depth_fr.txt",
"C5-nD_CCATGG_depth_fr.txt"
)
# Color plalette and other graphical parameters
# A (NT D nD) = red B (NT D nD) = green C (NT D nD) = blue
mypalette=c("red","red","red","#AAFF00","#AAFF00","#AAFF00","blue","blue","blue")
maxx=90
# ---------- FUNCTIONS ---------------
# returns tRNA sequence from coordinates and genome (5' to 3')
trnaseq5 <- function (genome, t) {
if (t$strand=="+"){strand="plus"}
if (t$strand=="-"){strand="minus"}
v=genome[t$start:t$end]
if (strand=="minus") {
v=reverseComplement(v)
}
return(as.character(v))
}
# compute jumps (ts) for 1 trna sequence
trnajump <- function (cov, from, to, strand) {
v=cov[from:to,strand]
if (strand=="plus") {
v=rev(v)
}
n=length(v)-1
vj=c(0) # sets first jump at 0
for (i in 1:n){
if (v[i]>99) {
if (v[i]<v[i+1]) {
jump=0
}else{
jump=(v[i]-v[i+1])/v[i]*100
}
vj=c(vj,jump)
} else {
vj=c(vj,NA)
}
}
if (length(vj)==0){vj=c(0)} # never return empty vector
return(vj)
}
# plot jumps for a list of tRNAs
# jmp1: array of jump values for drawing colored marks
# jmP2: vector of jump values for drawing bars
# tr: tRNA list
# genome: genome sequence
# allmods: strings of modified bases for all tRNAs
# pal: color palette for tick marks
plot_jumps <- function (jmp1, jmp2, tr, genome, allmods, pal, maxx) {
fname=paste0("jmp-", substr(tr[1,"name"],1,3),".pdf")
pdf(fname, paper="a4", width=8, height=11)
# fixed frame size
par(mfrow = c(6,1))
# par(mfrow = c(nrow(tr),1))
for (i in (1:nrow(tr))) {
t=tr[i,]
tname=as.character(t$name)
title=tr[i,"title"]
maxx=90
maxy=100
nbrep=length(allcov)
if (t$strand=="+"){strand="plus"}
if (t$strand=="-"){strand="minus"}
# compute jumps @ each position & replicates
jrep=c()
for (j in (1:nbrep)) {
cov=allcov[[j]]
jmp=rev(trnajump(cov,t$start,t$end,strand))
jmpl=length(jmp)
# padding to maxx
fill=rep(NA,max(0,maxx-jmpl))
jmp=c(fill, jmp)
print(length(jmp))
jrep=cbind(jrep, jmp)
}
# mean jump
meanj=apply(jrep,1,mean,na.rm=T)
seq=unlist(strsplit(trnaseq5(genome,t),split=''))
mods=unlist(allmods[tname])
# padd the sequence too
fill=rep(" ",max(0,maxx-(length(seq))))
seq=c(fill,seq)
mods=c(fill,mods)
par(mar=c(2, 2, 2, 0))
bp=barplot (meanj, border=NA, main=title, col="grey",ylim=c(0, maxy), xlim=c(0,110),axes=F)
# trick to allow all sequence labels to be displayed
impair=seq(1,length(seq),by=2)
pair=seq(0,length(seq),by=2)
axis(1, at=bp[impair], label=seq[impair], cex.axis=0.7, line = -1, tick=F)
axis(1, at=bp[pair], label=seq[pair], cex.axis=0.7, line = -1, tick=F)
axis(1, at=bp[impair], label=mods[impair], cex.axis=0.7, line = -0.4, tick=F, font=2)
axis(1, at=bp[pair], label=mods[pair], cex.axis=0.7, line = -0.4, tick=F, font=2)
yt=seq(0,maxy,by=10)
axis(2, cex.axis=0.7, at=yt, label=yt,line=-1)
for (j in (1:nbrep)) {
jmp=jrep[,j]
#points(x=bp, y=jmp, col=rainbow(nbrep)[j], pch="-", font=2)
points(x=bp, y=jmp, col=mypalette[j], pch="-", font=2)
}
}
dev.off()
}
# Compute trna jumps (ts) for tRNA list with replicates, 5' to 3'
# tr: a trna list
# allcov: array with all genome-wide coverages
# returns array with jumps for each replicate
# jumps are right justified as here (T=tRNA, J=jumps):
# <---------maxx-------->
# 5' --TTTTTTTTTTTTTTTTTTTTT 3'
# ---------JJJJJJJJJJJJJJ
trnajump_list <- function (allcov, tr, maxx) {
for (i in (1:nrow(tr))) {
t=tr[i,]
print (t$name)
nbrep=length(allcov)
if (t$strand=="+"){strand="plus"}
if (t$strand=="-"){strand="minus"}
# compute jumps @ each position & replicates
jrep=c()
for (j in (1:nbrep)) {
cov=allcov[[j]]
jmp=rev(trnajump(cov,t$start,t$end,strand))
jmpl=length(jmp)
# padding to maxx
fill=rep(NA,max(0,maxx-jmpl))
jmp=c(fill, jmp)
jrep=cbind(jrep, jmp)
}
return(jrep)
}
}
# mean jump
trnajump_mean <- function (allcov, tr, maxx) {
jrep<-trnajump_list (allcov, tr, maxx)
meanj=apply(jrep,1,mean,na.rm=T)
return(meanj)
}
# ------------------------
# MAIN
# ------------------------
#---------- read tRNA names and coordinates
alltrnas=read.table(trnacoordF, header=F, sep="\t")
trnanames=as.character(alltrnas$name)
colnames(alltrnas)=c("start","end","strand","name")
alltrnas=alltrnas[order(alltrnas$name),]
#---------- read genome
allChr<-readDNAStringSet(genomeF)
genome<-allChr[[1]]
genome=RNAString(genome)
#-------- read modomics strings
# beware: result is an array of lists
trnalines = readLines(modomicsF)
allmods<-c()
tnames<-c()
for (l in trnalines) {
print (nchar(l))
if (grepl (">", l)) {
tnames=c(tnames,substr(l,2,5))
}else{
# v<-unlist(strsplit(l,""))
v<-strsplit(l,"")
allmods <- cbind(allmods,v)
}
}
allmods=array(allmods)
rownames(allmods)=tnames
#-------- read coverage files
allcov <-list()
for (f in covF) {
f2=paste0(covdir,f)
cov=read.table(f2, header=F, sep="\t")
colnames(cov)=c("plus","minus")
allcov=append(allcov,list(cov))
print (nrow(cov))
}
names(allcov)=sub("(.*)\\..*", "\\1", covF) # keeps file prefix as lib name
# then for accessing one cov vector: allcov[[i]] (1 coverage track across whole genome)
# example: allcov[[1]][1:10,"minus"]
# read list of tRNAs to be printed
trnalines = readLines(trnaF)
# ------- Plot ts based on the sum of all coverages
printlist=c()
for (l in trnalines) {
v<-unlist(strsplit(l," "))
if (v[1]!="--") {
tr=alltrnas[grep(v[1], alltrnas$name),]
tr$title=l
printlist=rbind(printlist,tr)
} else {
print (printlist)
printlist=c()
}
}
# cimput sum of all coverages
covp=c()
covm=c()
nbrep=length(allcov)
for (i in (1:nbrep)) {
covp=cbind(covp,allcov[[i]][,"plus"])
covm=cbind(covm,allcov[[i]][,"minus"])
}
covp1=rowSums(covp)
covm1=rowSums(covm)
sumcov <- data.frame(covp1,covm1)
colnames(sumcov)=c("plus","minus")
#----------
trnajump_list(list(sumcov), alltrnas[1,], maxx)