Kostyrko Multiomics Analysis for Methylation EPIC and RNAseq

Description

This repository contains a set of R scripts specifically designed for the data analysis performed in Kostyrko et al. The repository provides a set of R scripts for analyzing Methylation EPIC and RNAseq (correlation and survival analysis) used in the original paper. This respository is a resource for researchers looking to replicate or build upon the findings of Kostyrko et al.

• PMID: 37407562 PMCID: PMC10322837 DOI: 10.1038/s41467-023-39591-2
• https://pubmed.ncbi.nlm.nih.gov/37407562/

Features

Methylation EPIC Analysis & RNAseq Correlation:

• This repo is tailored to handle Methylation EPIC data and RNAseq correlation analysis as performed in the Kostyrko et al study.
• Survival Analysis & Miscellaneous RNAseq Analysis:

Data

The dataset used for the Kostyrko et al study can be accessed from the GEO SuperSeries under the accession number GSE198289.

This SuperSeries is composed of the following SubSeries:

• GSE198289 UHRF1 is a mediator of KRAS driven oncogenesis in lung adenocarcinoma [RNA-seq]
• GSE198446 UHRF1 is a mediator of KRAS driven oncogenesis in lung adenocarcinoma [epic_methyl]
• GSE209923 UHRF1 is a mediator of KRAS driven oncogenesis in lung adenocarcinoma [shRNA]
• GSE233401 UHRF1 is a mediator of KRAS driven oncogenesis in lung adenocarcinoma [CRISPR_screen]

Minimum Requirements

• Download and install R from here. R (3.6.2)

• Download the dataset from GEO SuperSeries GSE198289.

• R packages

  • edgeR (3.28.1)
  • Minfi (1.32)
  • EpiDISH (2.2.2)
  • limma package(3.42.2)
  • missMethyl (1.20.4)
  • DMRcate (2.0.7)
  • survminer (0.4.6.999) and survival (3.1.11)
  • DGCA (1.0.2 )
  • methylGSA (1.4.9)

additional packages

Click to expand Bioinformatics and Genomics packages!

Genomic Data Annotation:

1. library(biomaRt): Tools for BioMart databases (like Ensembl).
2. library(BSgenome): Infrastructure for Bioconductor packages using large-scale genomic or other data.
3. library(org.Hs.eg.db): Mapping information for human genes.
4. library(GenomicFeatures): Tools for making and manipulating transcript centric annotations.
5. library(IlluminaHumanMethylation450kanno.ilmn12.hg19): Annotation data for the Illumina Human Methylation 450k array.
6. library(IlluminaHumanMethylationEPICanno.ilm10b4.hg19): Annotation data for the Illumina Human Methylation EPIC array.
7. library(IlluminaHumanMethylationEPICmanifest): Manifest file for Illumina's EPIC methylation arrays.
8. library(Homo.sapiens): Annotation data for the human genome.
9. library(rtracklayer): An interface to genome annotation files and the UCSC genome browser.

Genomic Data Analysis (Omics):

1. library(DESeq2): Differential gene expression analysis based on the negative binomial distribution.
2. library(edgeR): Empirical analysis of digital gene expression data in R.
3. library(GenomicRanges): Representations and manipulations of genomic intervals and variables defined along a genome.
4. library(GSVA): Gene set variation analysis for microarray and RNA-seq data.
5. library(Gviz): Plotting data and annotation information along genomic coordinates.
6. library(minfi): Tools to analyze Illumina's methylation arrays.
7. library(missMethyl): Analyzes differential methylation in the context of GC content.
8. library(methylGSA): Gene set testing for Illumina's methylation arrays.
9. library(pathview): Plots pathway maps and overlays experimental data.
10. library(sva): Surrogate Variable Analysis: identification and adjustment for hidden confounding factors.
11. library(biovizBase): Basic graphic utilities for visualization of genomic data.
12. library(ggbio): Visualization tools for genomic data.
13. library(limma): Linear models for microarray data.
14. library(pathfindR): An R package for comprehensive identification of enriched pathways in omics data through active subnetworks
15. library (DGCA): #Differential Gene Correlation Analysis

Genomic Data Analysis (Epigenetics):

1. library(EpiDISH): Epigenetic Dissection of Intra-Sample-Heterogeneity.
2. library(DMRcate): Detecting differentially methylated regions in CpG methylation data.

Heatmaps and Clustering:

1. library(clusterProfiler): Statistical analysis and visualization of functional profiles for genes and gene clusters.
2. library(ComplexHeatmap): Making complex heatmaps.
3. library(d3heatmap): Interactive heatmaps.
4. library(dendextend): Extending R's dendrogram functionality.
5. library(dendroextras): Extra functions to cut, label and colour dendrogram clusters.
6. library(parallelDist): Parallel distance matrix computation.

Visualization:

1. library(corrplot): Visualization of a correlation matrix.
2. library(factoextra): Extract and visualize the results of multivariate data analyses.
3. library(ggdendro): Create dendrograms using ggplot.
4. library(ggplot2): An implementation of the Grammar of Graphics.
5. library(ggplotify): Convert plot function call to 'ggplot' objects.
6. library(ggpubr): 'ggplot2' based publication ready plots.
7. library(ggpval): Annotate statistical significance onto 'ggplot' objects.
8. library(ggrepel): Automatically position non-overlapping text labels with 'ggplot2'.
9. library(gplots): Various R programming tools for plotting data.
10. library(gridExtra): Miscellaneous functions for "grid" graphics.
11. library(kableExtra): Build complex HTML or 'LaTeX' tables using 'kable()' and pipe syntax.
12. library(patchwork): The composer of ggplots.
13. library(RColorBrewer): ColorBrewer palettes.
14. library(VennDiagram): Generate high-resolution Venn and Euler plots.
15. library(Vennerable): Venn and Euler area-proportional diagrams.
16. library(wesanderson): Wes Anderson color palettes.
17. library(igraph): Network analysis and visualization.
18. library (ggbeeswarm) # Beeswarm plots helper
19. library(forestplot) # forest plot helper, mostly use in meta-analysis
20. library (ggridges) # Ridgeline plots
21. library(cowplot) # functions to align plots and arrange them into complex compound figures

Statistical Analysis:

1. library(FactoMineR): An R package for multivariate analysis.
2. library(fgsea): Fast gene set enrichment analysis.
3. library(MASS): Functions and datasets to support Venables and Ripley's MASS.
4. library(matrixStats): Functions that apply to rows and columns of matrices (and to vectors).
5. library(PerformanceAnalytics): Econometric tools for performance and risk analysis.
6. library(psych): Procedures for psychological, psychometric, and personality research.
7. library(survival): Survival analysis.
8. library(survminer): Drawing survival curves using 'ggplot2'.
9. library(vegan): Community Ecology Package.
10. library(scales): Scale functions for visualization.
11. library(Rtsne): T-distributed stochastic neighbor embedding using a Barnes-Hut implementation.
12. library(umap): Uniform Manifold Approximation and Projection.

Data Manipulation:

1. library(data.table): Extension of data.frame.
2. library(dplyr): A grammar of data manipulation.
3. library(DT): A wrapper of the JavaScript library 'DataTables'.
4. library(forcats): Tools for working with categorical variables (factors).
5. library(plyr): Tools for splitting, applying and combining data.
6. library(reshape): Flexibly reshape data.
7. library(stringr): Simple, consistent wrappers for common string operations.
8. library(tidyr): Easily tidy data with 'spread()' and 'gather()' functions.

Document Generation and Reporting:

1. library(knitr): A general-purpose tool for dynamic report generation in R.
2. library(pander): An R Pandoc writer.
3. library(stargazer) # LATEX, HTML and ASCII tables from R statistical output

File I/O:

1. library(openxlsx): Read, write and edit XLSX files.

Additional public datasets

LUAD, survival, correlations	RNAseq Counts	Broad GDAC ( Firehose )	https://gdac.broadinstitute.org/runs/stddata__2016_01_28/data/LUAD/20160128/gdac.broadinstitute.org_LUAD.Merge_rnaseqv2__illuminahiseq_rnaseqv2__unc_edu__Level_3__RSEM_genes__data.Level_3.2016012800.0.0.tar.gz
Pathway	database	MSigDB	https://www.gsea-msigdb.org/gsea/msigdb
EPIC annotation	Probe annotations	Bioconductor	https://bioconductor.org/packages/release/data/annotation/html/IlluminaHumanMethylationEPICanno.ilm10b4.hg19.html
TSG annotation	Annotations	UTHealth	https://bioinfo.uth.edu/TSGene/download.cgi?csrt=9609930864565957852
Probe Filter	Annotations	github	https://github.com/sirselim/illumina450k_filtering
ERV	Genomic annotation	UCSC genome	https://genome.ucsc.edu/cgi-bin/hgTables?hgta_doMainPage=1&hgta_group=rep&hgta_track=rmsk&hgta_table=rmsk
RNAseq index	Genomic index	GENCODE	ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_37/GRCh38.primary_assembly.genome.fa.gz
RNAseq GTF	Gene Annotation	GENCODE	ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_37/gencode.v37.annotation.gtf.gz