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--- a +++ b/6-Figure scripts/Fig 2.r @@ -0,0 +1,345 @@ +list.files() + +library(readxl) +excel_sheets("Fig 2 Source Data.xlsx") + +library(dplyr) + +# Figure 2a ------------------------------------------ + +# Figure 2a. metabolome ####### +rm(list = ls()) +dat.metab <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2a metabolome PCA") + +library(ggplot2) +pca <- dat.metab +Fig2a.metab.pca <- ggplot(pca,aes(PC1,PC2))+ + geom_point(size=2,aes(col=Group,shape=Cohort))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + scale_shape_manual(values=c(16,15)) + + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) +Fig2a.metab.pca + +pca$GROUP = paste(pca$Group,"|", pca$Cohort,sep = "") + +Fig2a.metab.pc1.density <- + ggplot(pca) + + geom_density(aes(x=PC1, group=GROUP, fill=Group, linetype=Cohort), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") +Fig2a.metab.pc1.density + +Fig2a.metab.pc2.density <- + ggplot(pca) + + geom_density(aes(x=PC2, group=GROUP, fill=Group, linetype=Cohort), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") + + coord_flip() +Fig2a.metab.pc2.density + + + +# Figure 2a. metabolome ####### +excel_sheets("Fig 2 Source Data.xlsx") +rm(list = ls()) +dat.trans <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2a transcriptome PCA") + +#library(ggplot2) +pca <- dat.trans +Fig2a.trans.pca <- ggplot(pca,aes(PC1,PC2))+ + geom_point(size=2,aes(col=Group,shape=Cohort))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + scale_shape_manual(values=c(16,15)) + + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) +Fig2a.trans.pca + +pca$GROUP = paste(pca$Group,"|", pca$Cohort,sep = "") + +Fig2a.trans.pc1.density <- + ggplot(pca) + + geom_density(aes(x=PC1, group=GROUP, fill=Group, linetype=Cohort), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") +Fig2a.trans.pc1.density + +Fig2a.trans.pc2.density <- + ggplot(pca) + + geom_density(aes(x=PC2, group=GROUP, fill=Group, linetype=Cohort), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") + + coord_flip() +Fig2a.trans.pc2.density + + +# Figure 2a. sputum proteome ####### +excel_sheets("Fig 2 Source Data.xlsx") +rm(list = ls()) +dat.sputum <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2a sputum proteome PCA") + +#library(ggplot2) +pca <- dat.sputum +Fig2a.sputum.pca <- ggplot(pca,aes(PC1,PC2))+ + geom_point(size=2,aes(col=Group))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + scale_shape_manual(values=c(16,15)) + + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) +Fig2a.sputum.pca + +Fig2a.sputum.pc1.density <- + ggplot(pca) + + geom_density(aes(x=PC1, group=Group, fill=Group), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") +Fig2a.sputum.pc1.density + +Fig2a.sputum.pc2.density <- + ggplot(pca) + + geom_density(aes(x=PC2, group=Group, fill=Group), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") + + coord_flip() +Fig2a.sputum.pc2.density + + +# Figure 2a. serum proteome ####### +excel_sheets("Fig 2 Source Data.xlsx") +rm(list = ls()) +dat.serum <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2a serum proteome PCA") + +#library(ggplot2) +pca <- dat.serum +Fig2a.serum.pca <- ggplot(pca,aes(PC1,PC2))+ + geom_point(size=2,aes(col=Group))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + scale_shape_manual(values=c(16,15)) + + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) +Fig2a.serum.pca + +Fig2a.serum.pc1.density <- + ggplot(pca) + + geom_density(aes(x=PC1, group=Group, fill=Group), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") +Fig2a.serum.pc1.density + +Fig2a.serum.pc2.density <- + ggplot(pca) + + geom_density(aes(x=PC2, group=Group, fill=Group), + color="black", alpha=0.6, position = 'identity') + + scale_fill_manual(values=c("#f0999f","#46bbc0")) + + theme_bw() + + scale_linetype_manual(values = c("solid","dashed"))+ + labs(fill="") + + coord_flip() +Fig2a.serum.pc2.density + + +# Figure 2b ------------------------------------------ +excel_sheets("Fig 2 Source Data.xlsx") +rm(list = ls()) +metab.modules <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2b metab modules") +metab.features <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2b metab features") + +# Figure 2b.density of modules #### +data <- metab.modules +library(reshape2) +data2<-melt(data,id.vals=c("NAME","CLASS","GROUP") ) +md.levels <- c("MEbrown4","MEmediumpurple1","MEskyblue3","MElavenderblush3","MEslateblue", + "MEsalmon2","MEbisque4","MEplum","MEmidnightblue","MEindianred4") + +data2$variable=factor(data2$variable,levels=md.levels) +data2$GROUP4 <- paste(data2$GROUP,data2$CLASS, sep = "|") +Fig2b.density <- ggplot(data2,aes(data2$value))+ + geom_density(aes(group=data2$GROUP4,fill=data2$CLASS,alpha=0.4,linetype=data2$GROUP))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + facet_wrap(~data2$variable,scale="free",ncol=1)+ + scale_linetype_manual(values=c("solid","dashed"))+ + scale_x_continuous(limits=c(-0.2,0.2)) + + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) +Fig2b.density +ggsave(Fig2b.density, filename = "Fig2b.density.pdf",device = "pdf", width = 4, height = 10) + +# Figure 2b.heatmap #### +data <- metab.features +data.Sz <- metab.features %>% select(starts_with("Z")) +data.Gz <- metab.features %>% select(!starts_with("Z")) %>% select(-Metabolite, -Group) + +colors<-c(seq(-4,0,length=51),seq(0.1,4,length=50)) +my_palette<- colorRampPalette(c("#EE9494","#F6E572","#94EE9D","#94E1EE","#94B3EE","white","white","white","white","white","white","white"))(n=100) +library(gplots) +pdf( "Fig2b.metab.hm.Sz.pdf") #save figure2b.metab.heamap.Shenzhen in pdf +heatmap.2(as.matrix(data.Sz), + breaks=colors, scale="row", trace="none", + Rowv=FALSE, dendrogram='column',hclustfun = function(x) hclust(x, method="ward.D2"), + col=rev(my_palette)) +dev.off() + +pdf( "Fig2b.metab.hm.Gz.pdf") #save figure2b.metab.heamap.Guangzhou in pdf +heatmap.2(as.matrix(data.Gz), + breaks=colors, scale="row", trace="none", + Rowv=FALSE, dendrogram='column',hclustfun = function(x) hclust(x, method="ward.D2"), + col=rev(my_palette)) +dev.off() + + +## Figure 2c ------------------------------------------ +excel_sheets("Fig 2 Source Data.xlsx") +rm(list = ls()) +trans.modules <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2c trans modules") +trans.features <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2c trans features") + +# Figure 2c.density of modules #### +data <- trans.modules +#library(reshape2) +data2<-melt(data,id.vals=c("NAME","CLASS","GROUP") ) +md.levels <- c("MEcyan","MEdarkmagenta.1", + "MEorangered4","MEskyblue2", + "MEblueviolet","MEthistle4", + "MElightyellow","MEchocolate4", + "MEbisque4.1","MEsnow4") + +data2$variable=factor(data2$variable,levels=md.levels) +data2$GROUP4 <- paste(data2$GROUP,data2$CLASS, sep = "|") +Fig2c.density <- ggplot(data2,aes(value))+ + geom_density(aes(group=GROUP4,fill=CLASS,alpha=0.4,linetype=GROUP))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + facet_wrap(~variable,scale="free",ncol=1)+ + scale_linetype_manual(values=c("solid","dashed"))+ + scale_x_continuous(limits=c(-0.2,0.2)) + + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) +Fig2c.density +ggsave(Fig2c.density, filename = "Fig2c.density.pdf",device = "pdf", width = 4, height = 10) + + +# Figure 2c.heatmap #### +data <- trans.features +data.Sz <- trans.features %>% select(starts_with("Z")) +data.Gz <- trans.features %>% select(!starts_with("Z")) %>% select(-Gene, -Module) + +colors <- c(seq(-4, 0, length=51), seq(0.1, 4, length=50)) +my_palette<- colorRampPalette(c("#EE9494","#F6E572","#94EE9D","#94E1EE","#94B3EE","white","white","white","white","white","white","white"))(n=100) +#library(gplots) +#pdf( "Fig2c.trans.hm.Sz.pdf") #save figure2c.trans.heamap.Shenzhen in pdf +heatmap.2(as.matrix(data.Sz), + breaks=colors, scale="row", trace="none", + Rowv=FALSE, dendrogram='column',hclustfun = function(x) hclust(x, method="ward.D2"), + col=rev(my_palette)) +#dev.off() + +pdf( "Fig2c.trans.hm.Gz.pdf") #save figure2c.trans.heamap.Guangzhou in pdf +heatmap.2(as.matrix(data.Gz), + breaks=colors, scale="row", trace="none", hclustfun = function(x) hclust(x, method="ward.D2"), + Rowv=FALSE, dendrogram='column', + col=rev(my_palette)) +dev.off() + + +## Figure 2d ------------------------------------------ +excel_sheets("Fig 2 data.xlsx") +rm(list = ls()) +prot.features <- read_excel("Fig 2 Source Data.xlsx", sheet = "Fig 2d proteome features") +sputum.p <- prot.features %>% select(SampleID, Group, starts_with("Sputum")) +serum.p <- prot.features %>% select(SampleID, Group, starts_with("serum")) + +# Figure2d. sputum proteome #### +data <- sputum.p + +data.l <- reshape2::melt(data, id.vars=c("SampleID", "Group")) +sapply(data.l,class) +data.l_positive <- data.l %>% filter(value > 0) + +H_median_df <- data.l_positive %>% filter(Group == "H") %>% + group_by(variable) %>% summarise(H_median =median(value)) %>% + as.data.frame(stringsAsFactors=F) + +plotDat <- + merge(data.l_positive,H_median_df,by="variable") %>% + mutate(normalized_value = value/H_median) %>% + mutate(normalized.log = log10(normalized_value)) + + +Fig2d.sputum <- ggplot(plotDat,aes(normalized.log))+ + geom_density(aes(group=Group,fill=Group,alpha=0.4))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + facet_wrap(~variable,scale="free",ncol = 1) + + # scale_linetype_manual(values=c("solid","dashed"))+ + #scale_x_continuous(limits=c(-0.2,0.2))+ + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) + +Fig2d.sputum +ggsave(Fig2d.sputum, filename = 'Fig2d.sputum.pdf', device = "pdf",width = 4,height = 10) + + + +# Figure2d. serum proteome #### +data <- serum.p + +data.l <- reshape2::melt(data, id.vars=c("SampleID", "Group")) +sapply(data.l,class) +data.l_positive <- data.l %>% filter(value > 0) + +H_median_df <- data.l_positive %>% filter(Group == "H") %>% + group_by(variable) %>% summarise(H_median =median(value)) %>% + as.data.frame(stringsAsFactors=F) + +plotDat <- + merge(data.l_positive,H_median_df,by="variable") %>% + mutate(normalized_value = value/H_median) %>% + mutate(normalized.log = log10(normalized_value)) + + +Fig2d.serum <- ggplot(plotDat,aes(normalized.log))+ + geom_density(aes(group=Group,fill=Group,alpha=0.4))+ + scale_color_manual(values=c("#f0999f","#46bbc0"))+ + facet_wrap(~variable,scale="free",ncol = 1) + + # scale_linetype_manual(values=c("solid","dashed"))+ + scale_x_continuous(limits=c(-0.2,0.2))+ + theme_bw()+theme(axis.line = element_line(colour = "black"), + panel.grid.major = element_blank(), + panel.grid.minor = element_blank(), + panel.background = element_blank()) + +Fig2d.serum +ggsave(Fig2d.serum, filename = 'Fig2d.serum.pdf', device = "pdf",width = 4,height = 4)