Graphics issue in plotFusion and plotFusionReads

[komalsrathi]

Hi Stian,

I am facing a lot of graphics issue so for simplicity, I am just using your test code so it is easier for you to reproduce the problem:

library(chimeraviz)

# Get reference to results file from deFuse
defuse833ke <- system.file(
  "extdata",
  "defuse_833ke_results.filtered.tsv",
  package="chimeraviz")

# Load the results file into a list of fusion objects
fusions <- importDefuse(defuse833ke, "hg19", 1)

length(fusions)

fusion <- getFusionById(fusions, 5267)

fastq1 <- system.file(
  "extdata",
  "reads_supporting_defuse_fusion_5267.1.fq",
  package = "chimeraviz")

fastq2 <- system.file(
  "extdata",
  "reads_supporting_defuse_fusion_5267.2.fq",
  package = "chimeraviz")

referenceFilename <- "reference.fa"
writeFusionReference(fusion = fusion, filename = referenceFilename)
source(system.file(
  "scripts",
  "rsubread.R",
  package="chimeraviz"))

# Then create index
rsubreadIndex(referenceFasta = referenceFilename)
rsubreadAlign(
  referenceName = referenceFilename,
  fastq1 = fastq1,
  fastq2 = fastq2,
  outputBamFilename = "fusionAlignment")

if(!exists("bamfile5267")) {
  bamfile5267 <- system.file(
    "extdata",
    "5267readsAligned.bam",
    package="chimeraviz")
}

fusion <- addFusionReadsAlignment(fusion, bamfile5267)
plotFusionReads(fusion = fusion)

# another bam file
if(!exists("bamfile5267")) {
  bamfile5267 <- system.file(
    "extdata",
    "5267readsAligned.bam",
    package="chimeraviz")
}
fusion <- addFusionReadsAlignment(fusion, bamfile5267)
plotFusionReads(fusion = fusion, showAllNucleotides = TRUE)

At this point, this is the plot I am getting -

Why is it not showing the ATGC and showing colored bars instead?

Another plot that is not showing correctly:

if(!exists("defuse833ke"))
  defuse833ke <- system.file(
    "extdata",
    "defuse_833ke_results.filtered.tsv",
    package = "chimeraviz")

# Then load the fusion events
fusions <- importDefuse(defuse833ke, "hg19", 1)
fusion <- getFusionById(fusions, 5267)
if(!exists("edbSqliteFile"))
  edbSqliteFile <- system.file(
    "extdata",
    "Homo_sapiens.GRCh37.74.sqlite",
    package="chimeraviz")
# Then load it
edb <- ensembldb::EnsDb(edbSqliteFile)

if(!exists("fusion5267and11759reads"))
  fusion5267and11759reads <- system.file(
    "extdata",
    "fusion5267and11759reads.bam",
    package = "chimeraviz")

plotFusion(
  fusion = fusion,
  bamfile = bamfile5267,
  edb = edb,
  nonUCSC = TRUE,
  reduceTranscripts = TRUE)

Here the red arc is not showing properly that should connect the two exons of RCC1 and HENMT1.

Here is my sessionInfo:

> sessionInfo()
R version 3.4.2 (2017-09-28)
Platform: x86_64-apple-darwin15.6.0 (64-bit)
Running under: OS X El Capitan 10.11.6

Matrix products: default
BLAS: /System/Library/Frameworks/Accelerate.framework/Versions/A/Frameworks/vecLib.framework/Versions/A/libBLAS.dylib
LAPACK: /Library/Frameworks/R.framework/Versions/3.4/Resources/lib/libRlapack.dylib

locale:
[1] en_US.UTF-8

attached base packages:
 [1] grid      stats4    parallel  stats     graphics  grDevices utils     datasets  methods   base     

other attached packages:
 [1] chimeraviz_1.2.2       ensembldb_2.0.4        AnnotationFilter_1.0.0 GenomicFeatures_1.28.5 AnnotationDbi_1.38.2  
 [6] Biobase_2.36.2         Gviz_1.20.0            GenomicRanges_1.28.6   GenomeInfoDb_1.12.3    Biostrings_2.44.2     
[11] XVector_0.16.0         IRanges_2.10.5         S4Vectors_0.14.7       BiocGenerics_0.22.1    BiocInstaller_1.26.1  

loaded via a namespace (and not attached):
 [1] ProtGenerics_1.8.0            bitops_1.0-6                  matrixStats_0.52.2           
 [4] bit64_0.9-7                   RColorBrewer_1.1-2            httr_1.3.1                   
 [7] rprojroot_1.2                 tools_3.4.2                   backports_1.1.1              
[10] R6_2.2.2                      DT_0.2                        rpart_4.1-11                 
[13] Hmisc_4.0-3                   DBI_0.7                       lazyeval_0.2.0               
[16] colorspace_1.3-2              nnet_7.3-12                   gridExtra_2.3                
[19] bit_1.1-12                    curl_3.0                      compiler_3.4.2               
[22] htmlTable_1.9                 DelayedArray_0.2.7            rtracklayer_1.36.6           
[25] scales_0.5.0                  checkmate_1.8.4               readr_1.1.1                  
[28] RCircos_1.2.0                 stringr_1.2.0                 digest_0.6.12                
[31] Rsamtools_1.28.0              foreign_0.8-69                rmarkdown_1.6                
[34] Rsubread_1.26.1               base64enc_0.1-3               dichromat_2.0-0              
[37] pkgconfig_2.0.1               htmltools_0.3.6               BSgenome_1.44.2              
[40] htmlwidgets_0.9               rlang_0.1.2                   RSQLite_2.0                  
[43] shiny_1.0.5                   BiocParallel_1.10.1           acepack_1.4.1                
[46] VariantAnnotation_1.22.3      RCurl_1.95-4.8                magrittr_1.5                 
[49] GenomeInfoDbData_0.99.0       Formula_1.2-2                 Matrix_1.2-11                
[52] Rcpp_0.12.13                  munsell_0.4.3                 stringi_1.1.5                
[55] yaml_2.1.14                   SummarizedExperiment_1.6.5    zlibbioc_1.22.0              
[58] org.Hs.eg.db_3.4.1            plyr_1.8.4                    AnnotationHub_2.8.2          
[61] blob_1.1.0                    lattice_0.20-35               splines_3.4.2                
[64] hms_0.3                       knitr_1.17                    biomaRt_2.32.1               
[67] XML_3.98-1.9                  evaluate_0.10.1               biovizBase_1.24.0            
[70] latticeExtra_0.6-28           data.table_1.10.4-1           httpuv_1.3.5                 
[73] gtable_0.2.0                  ggplot2_2.2.1                 mime_0.5                     
[76] xtable_1.8-2                  ArgumentCheck_0.10.2          survival_2.41-3              
[79] tibble_1.3.4                  GenomicAlignments_1.12.2      memoise_1.1.0                
[82] cluster_2.0.6                 interactiveDisplayBase_1.14.0 BiocStyle_2.4.1      

[stianlagstad]

The first issue is that there's not enough rom on the screen to actually show you all the nucleotides. You can limit the amount of nucleotides with the nucleotideAmount parameter - that'll let you experiment a bit and find the limit. Try setting it to 10 to start, and also set showAllNucleotides to FALSE.

plotFusionReads(fusion = fusion, showAllNucleotides = FALSE, nucleotideAmount = 10)

---

I reproduced the second issue this morning on a mac at work. For some reason it draws the arc outside of the actual plot. But it seems to be a mac-specific problem. I'm not able to reproduce the error with my home laptop running Ubuntu. This code:

library(chimeraviz)
if(!exists("defuse833ke"))
  defuse833ke <- system.file(
    "extdata",
    "defuse_833ke_results.filtered.tsv",
    package = "chimeraviz")

# Then load the fusion events
fusions <- importDefuse(defuse833ke, "hg19", 1)
fusion <- getFusionById(fusions, 5267)
if(!exists("edbSqliteFile"))
  edbSqliteFile <- system.file(
    "extdata",
    "Homo_sapiens.GRCh37.74.sqlite",
    package="chimeraviz")
# Then load it
edb <- ensembldb::EnsDb(edbSqliteFile)

if(!exists("fusion5267and11759reads"))
  fusion5267and11759reads <- system.file(
    "extdata",
    "fusion5267and11759reads.bam",
    package = "chimeraviz")

if(!exists("bamfile5267")) {
  bamfile5267 <- system.file(
    "extdata",
    "5267readsAligned.bam",
    package="chimeraviz")
}

plotFusion(
  fusion = fusion,
  bamfile = bamfile5267,
  edb = edb,
  nonUCSC = TRUE,
  reduceTranscripts = TRUE)

Produces this plot for me:

Edit: Back on the mac, that same code produces this plot:

I'll look into this as soon as I can!

[stianlagstad]

Just to show a sign of life: So far I'm clueless. In the plotFusion function I use Gviz::coords() to fetch the coordinates of where the exons are plotted in the picture. I use these to decide where to draw the red arc. For some reason the y coordinates those code lines returns on a mac are off. I don't know why yet. I don't really think it's a Gviz issue.

Relevant chimeraviz code from plotFusion.R:

    # Get coordinates for the exons in the plot
    exonCoordinates1 <- Gviz::coords(res1$GeneRegionTrack)
    exonCoordinates2 <- Gviz::coords(res2$GeneRegionTrack)

[stianlagstad]

After getting some help on the bioc-devel list I got a bit further. The arc turns up where it's supposed to when I use another graphics device. Could you try that as well? I.e. try:

# Load data and example fusion event
defuse833ke <- system.file(
  "extdata",
  "defuse_833ke_results.filtered.tsv",
  package="chimeraviz")
fusions <- importDefuse(defuse833ke, "hg19", 1)
fusion <- getFusionById(fusions, 5267)
# Load edb
edbSqliteFile <- system.file(
  "extdata",
  "Homo_sapiens.GRCh37.74.sqlite",
  package="chimeraviz")
edb <- ensembldb::EnsDb(edbSqliteFile)
# bamfile with reads in the regions of this fusion event
bamfile5267 <- system.file(
  "extdata",
  "fusion5267and11759reads.bam",
  package="chimeraviz")
# Temporary file to store the plot
pngFilename <- "testplot.png"
# Open device
png(pngFilename, width = 1000, height = 750)
# Plot!
plotFusion(
  fusion = fusion,
  bamfile = bamfile5267,
  edb = edb,
  nonUCSC = TRUE)
# Close device
dev.off()

and then see what testplot.png looks like?

[komalsrathi]

Yes this works for me -- thanks! I am testing the entire code now and will update you on it.

Thanks a lot!

[stianlagstad]

Great! I'll look more into the default graphics driver on mac and try to figure out the issue. But I'll close this :)