Volcano plot not centered at origin
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Entering edit mode
12 weeks ago
mm2568 • 0

I am making a volcano plot for my phosphoproteomics data and seemingly the data are not centered around the origin. The reason that I know there is something wrong is because before my code was producing the correct volcano plot, and now it is shifted. What are some reasons why it could be shifted without showing all my code? The only difference between the two is that I didn't filter out as many phosphopeptides in the second plot. Any obvious red flags that I am not seeing?

I am making the plot in Enhaced Volcano in R with this code:

Rap_nutri_df <- cbind(final_phosphoproteomics_nutri_caltech$Phosphorylation_Site,
                  final_phosphoproteomics_nutri_caltech$`Log2(Ratio_Rap_v_Control)`, #this is shifted!!!!
                  final_phosphoproteomics_nutri_caltech$`Adj_p-val_Rap_v_Control`)
colnames(Rap_nutri_df) <- c("phosphopeptide", "log2FoldChange","adj_pvalue")

R_nutri_df <- transform(R_nutri_df, log2FoldChange = as.numeric(log2FoldChange), 
                          adj_pvalue = as.numeric(adj_pvalue),
                          phosphopeptide = as.character(phosphopeptide))
options(ggrepel.max.overlaps = Inf)
EnhancedVolcano(R_nutri_df, 
                lab = '', #R_nutri_df$phosphopeptide,
                x = 'log2FoldChange',
                y = 'adj_pvalue',
                title = 'R vs Control',
                labSize = 1.0,
                pCutoff = 0.05,
                FCcutoff = 1.5,
                legendPosition = 'top',
                legendLabSize = 12,
                legendIconSize = 4.0,
                col=c('black', 'black', 'black', 'green1'),
                xlim = c(-5, 5),
                gridlines.major = TRUE,
                gridlines.minor = TRUE,
                #drawConnectors = TRUE, #draw arrows from data points
                widthConnectors = 0.75,
                shape=15) + #square shape of data points
  ggplot2::coord_cartesian(xlim=c(-3.1, 3.1))

enter image description here

enter image description here

R • 218 views
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3
Entering edit mode

This is a DESeq2/edgeR/whatever you're using upstream question, not an EnhancedVolcano question. Something upstream is causing the skew, so I'd recommend taking a close look at that and utilizing MAplots to determine how exactly it's happening.

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0
Entering edit mode

It's hard to pinpoint anything, but it's not the plotting function. However, you show us this

Rap_nutri_df <- 

but then you plot this

R_nutri_df <- transform(R_nutri_df, ...)

so where's the Rap? Maybe you already had a R_nutri_df object in your workspace.

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