Abstract RNA sequencing of time-course experiments results in three-way count data where the dimensions are the genes, the time points and the biological units. Clustering RNA-seq data allows to extract groups of co-expressed genes over time. After standardisation, the normalised counts of individual genes across time points and biological units have similar properties as compositional data. We propose the following procedure to suitably cluster three-way RNA-seq data: (1) pre-process the RNA-seq data by calculating the normalised expression profiles, (2) transform the data using the additive log ratio transform to map the composition in the D-part Aitchison simplex to a $$D-1$$ D - 1 -dimensional Euclidean vector, (3) cluster the transformed RNA-seq data using matrix-variate Gaussian mixture models and (4) assess the quality of the overall cluster solution and of individual clusters based on cluster separation in the transformed space using density-based silhouette information and on compactness of the cluster in the original space using cluster maps as a suitable visualisation. The proposed procedure is illustrated on RNA-seq data from fission yeast and results are also compared to an analogous two-way approach after flattening out the biological units.

UNLABELLED: A model class of finite mixtures of linear additive models is presented. The component-specific parameters in the regression models are estimated using regularized likelihood methods. The advantages of the regularization are that (i) the pre-specified maximum degrees of freedom for the splines is less crucial than for unregularized estimation and that (ii) for each component individually a suitable degree of freedom is selected in an automatic way. The performance is evaluated in a simulation study with artificial data as well as on a yeast cell cycle dataset of gene expression levels over time. AVAILABILITY: The latest release version of the R package flexmix is available from CRAN (http://cran.r-project.org/).

SUMMARY: Finite mixture models are routinely applied to time course microarray data. Due to the complexity and size of this type of data, the choice of good starting values plays an important role. So far initialization strategies have only been investigated for data from a mixture of multivariate normal distributions. In this work several initialization procedures are evaluated for mixtures of regression models with and without random effects in an extensive simulation study on different artificial datasets. Finally, these procedures are also applied to a real dataset from Escherichia coli. AVAILABILITY: The latest release versions of R packages flexmix, gcExplorer and kernlab are always available from CRAN (http://cran.r-project.org/). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Abstract Summary: Cluster analysis plays an important role in the analysis of gene expression data since the early beginning of microarray studies and is routinely used to find groups of genes with common expression pattern. In order to make cluster analysis helpful for users, visualization of cluster solutions is of utmost importance. Here, we present the new R package gcExplorer for the interactive exploration of gene clusters. gcExplorer facilitates the interpretation of cluster results and allows to investigate extensive information about clusters. Availability: The latest release of gcExplorer is always available at the Comprehensive R Archive Network CRAN: http://cran.R-project.org/package=gcExplorer. See the README file in the package for detailed installation instructions. Contact: [email protected] Supplementary information: Supplementary data are available at Bioinformatics online.

BACKGROUND: Many different cluster methods are frequently used in gene expression data analysis to find groups of co-expressed genes. However, cluster algorithms with the ability to visualize the resulting clusters are usually preferred. The visualization of gene clusters gives practitioners an understanding of the cluster structure of their data and makes it easier to interpret the cluster results. RESULTS: In this paper recent extensions of R package gcExplorer are presented. gcExplorer is an interactive visualization toolbox for the investigation of the overall cluster structure as well as single clusters. The different visualization options including arbitrary node and panel functions are described in detail. Finally the toolbox can be used to investigate the quality of a given clustering graphically as well as theoretically by testing the association between a partition and a functional group under study. CONCLUSION: It is shown that gcExplorer is a very helpful tool for a general exploration of microarray experiments. The identification of potentially interesting gene candidates or functional groups is substantially accelerated and eased. Inferential analysis on a cluster solution is used to judge its ability to provide insight into the underlying mechanistic biology of the experiment.