csSAM package - Cell Type-Specific Statistical Analysis of Microarray
Tissues are often made up of multiple cell-types. Each with its own functional attributes and molecular signature. Yet, the proportions of any given cell-type in a sample can vary markedly. This results in a significant loss of sensitivity in gene expression studies and great difficulty in identifying the cellular source of any perturbations. Here we present a statistical methodology (cell-type specific Significance Analysis of Microarrays or csSAM) which, given microarray data from two groups of biological samples and the relative cell-type frequencies of each sample, estimates in a virtual manner the gene expression data for each cell-type at a group level, and uses these to identify differentially expressed genes at a cell-type specific level between groups.
The lower limit for the number of samples needed for deconvolving the cell-specific expression of N cell-types is N+1. For a single colour array - the result could be interpreted as the average expression level of a given gene in a cell-type of that group. Multiplied by the frequency of a given cell-type in an individual in the group, it is the amount contributed by that cell type to the overall measured expression on the array.
csSamWrapper - Single wrapper function performs all
functionality. csfit: For deconvolving the average cell-type specific
expression for each cell-type in a given group.
csSAMfit - For fitting csSAM model on Biobase::ExpressionSet objects
with the convenience of a formula interface.
csSAM - For calculating the
contrast between every pair of cells being compared between the two
fdrCsSAM - Estimate the false discovery rate for each cell-type
findSigGene - Identifies the list of differentially
expressed genes in a given cell-type at a given FDR cutoff.
plotCsSAM - Plots a fdr plot of their results.
Additional functions exists (e.g.,
to contrast csSAM with the tissue heterogeneity ignorant SAM).
Shen-Orr SS, Tibshirani R, Khatri P, Bodian DL, Staedtler F, Perry NM, Hastie T, Sarwal MM, Davis MM and Butte AJ (2010). “Cell type-specific gene expression differences in complex tissues.” Nature methods, 7(4), pp. 287-9. ISSN 1548-7105, doi: 10.1038/nmeth.1439 (URL: http://doi.org/10.1038/nmeth.1439).
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library("csSAM") ## ## Generate random dataset ## set.seed(143) k <- 5 # number of cell types ng <- 500 # number of genes p <- 20 # number of samples ndiff <- 100 # number of genes differentially expressed # true cell-specific signatures H1 <- matrix(rnorm(5*ng), ncol=ng) H2 <- H1 # create differential expression for 3rd cell type H2[3,1:ndiff] <- H2[3,1:ndiff] + 5 # cell frequency matrix per sample cc <- matrix(runif(p*k), ncol=k) cc <- t(scale(t(cc), center=FALSE, scale=rowSums(cc))) colnames(cc) <- paste('cellType', 1:ncol(cc), sep="") # global expression matrix G <- rbind(cc[1:10, ] %*% H1, cc[11:p, ] %*%H2 ) + matrix(rnorm(p*ng), ncol=ng) # sample classes (2 groups) y <- gl(2, p/2) fileName = "Example File.pdf"; # Now run, either using the wrapper # NB: more permutations would be needed for real data deconvResults = csSamWrapper(G, cc, y, nperms = 50, alternative = "two.sided" , standardize = TRUE , medianCenter = TRUE , fileName = fileName) # Or by calling each function independently: # this is useful if you want to perform only cell-specific expression # without differential expression. # run csSAM fit <- csSAMfit(t(G) ~ y | cc, nperms = 200, verbose = TRUE, .rng = 123) top <- csTopTable(fit, alternvative = 'two.sided') head(top[]) # run SAM tt.sam <- runSAM(G, y) falseDiscovRSAM <- fdrSAM(G, y, tt.sam, nperms=200, alternative = 'two.sided') plotCsSAM(fit, falseDiscovRSAM, alternative='two.sided', fileName)
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