Analysis and design of RNA sequencing experiments for identifying isoform regulation

By Y. Katz, Eric T. Wang, Edoardo M. Airoldi, C. Burge. Nature Methods, 2010

1,367 citations recorded by the source corpus.

Through alternative splicing, most human genes express multiple isoforms that often differ in function. To infer isoform regulation from high-throughput sequencing of cDNA fragments (RNA-seq), we developed the mixture-of-isoforms (MISO) model, a statistical model that estimates expression of alternatively spliced exons and isoforms and assesses confidence in these estimates. Incorporation of mRNA fragment length distribution in paired-end RNA-seq greatly improved estimation of alternative-splicing levels. MISO also detects differentially regulated exons or isoforms. Application of MISO implicated the RNA splicing factor hnRNP H1 in the regulation of alternative cleavage and polyadenylation, a role that was supported by UV cross-linking–immunoprecipitation sequencing (CLIP-seq) analysis in human cells. Our results provide a probabilistic framework for RNA-seq analysis, give functional insights into pre-mRNA processing and yield guidelines for the optimal design of RNA-seq experiments for studies of gene and isoform expression.

Subfields: Molecular Biology

View Paper: https://www.semanticscholar.org/paper/1826051b68883729f54fdaaa33d0f557209286ea

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Citations in the corpus

TCC: an R package for comparing tag count data with robust normalization strategies (2013, methodology): “In addition to the RDR6 knockout dataset, we also analyze four other experimental datasets (called “ sultan ” [4], “ gilad ” [22], “ maqc ” [23], and “ katz.mouse ” [24]) obtained from the ReCount database [25].”
Adult Mouse Cortical Cell Taxonomy by Single Cell Transcriptomics (2016, positive): “Because MISO does not accommodate replicates, to calculate MISO Ψ , we pooled 10 randomly selected single cell samples for each cell type (20 for broad glutamatergic, GABAergic and non-neuronal types) for each pairwise comparison.”
Differential gene and transcript expression analysis of RNA-seq experiments with TopHat and Cufflinks (2012, background): “The packages are ‘read-only’ and do not need to be redownloaded with each run of the protocol.”
YTHDC1 mediates nuclear export of N6-methyladenosine methylated mRNAs (2017, methodology): “Analysis using Mixture of Isoforms (MISO) software (Katz et al., 2010) identified 426 alternative splicing events with a difference in percent spliced in (psi, ) value of 0.2 or greater (Figure 4—figure supplement Figure 2 continued DOI: https://doi.org/10.7554/eLife.31311.008 Figure supplement 1.”
Extensive cross-regulation of post-transcriptional regulatory networks in Drosophila (2015, positive): “Other proteins that have been shown to have pleiotropic effects include NOVA2 (Licatalosi et al. 2008), MBNL (Han et al. 2010) family proteins, and HNRNPH1 (Katz et al. 2010).”
Single-cell mRNA quantification and differential analysis with Census (2017, methodology): “The Dirichlet has proven effective in previous analyses of splicing changes in bulk RNA-seq studies 51 .”
MATS: a Bayesian framework for flexible detection of differential alternative splicing from RNA-Seq data (2012, methodology): “This is different from and more general than the independent uniform priors used by previous methods (11,16).”
Mechanisms and consequences of alternative polyadenylation (2011, positive): “Specifically, Katz et al. (2010) used a statistical model to infer isoform regulation from RNA-seq data.”