Dataset Information

Long-read transcriptome sequencing reveals abundant promoter diversity in distinct molecular subtypes of gastric cancer

ABSTRACT: Deregulated gene expression is a hallmark of cancer, however most studies to date have analyzed short-read RNA-sequencing data with inherent limitations. Here, we combine PacBio long-read isoform sequencing (Iso-Seq) and Illumina paired-end short read RNA sequencing to comprehensively survey the transcriptome of gastric cancer (GC), a leading cause of global cancer mortality. We performed full-length transcriptome analysis across 10 GC cell lines covering four major GC molecular subtypes (chromosomal unstable, Epstein-Barr positive, genome stable and microsatellite unstable). We identify 60,239 non-redundant full-length transcripts, of which >66% are novel compared to current transcriptome databases. Novel isoforms are more likely to be cell-line and subtype specific, expressed at lower levels with larger number of exons, with longer isoform/coding sequence lengths. Most novel isoforms utilize an alternate first exon, and compared to other alternative splicing categories are expressed at higher levels and exhibit higher variability. Collectively, we observe alternate promoter usage in 25% of detected genes, with the majority (84.2%) of known/novel promoter pairs exhibiting potential changes in their coding sequences. Mapping these alternate promoters to TCGA GC samples, we identify several cancer-associated isoforms, including novel variants of oncogenes. Tumor-specific transcript isoforms tend to alter protein coding sequences to a larger extent than other isoforms. Analysis of outcome data suggests that novel isoforms may impart additional prognostic information. Our results provide a rich resource of full-length transcriptome data for deeper studies of GC and other gastrointestinal malignancies.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Stomach

DISEASE(S): Stomach Cancer

SUBMITTER: Dennis Kappei

LAB HEAD: Dennis Kappei

PROVIDER: PXD023373 | Pride | 2021-02-02

REPOSITORIES: Pride

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Publications

Long-read transcriptome sequencing reveals abundant promoter diversity in distinct molecular subtypes of gastric cancer.

Huang Kie Kyon KK Huang Jiawen J Wu Jeanie Kar Leng JKL Lee Minghui M Tay Su Ting ST Kumar Vikrant V Ramnarayanan Kalpana K Padmanabhan Nisha N Xu Chang C Tan Angie Lay Keng ALK Chan Charlene C Kappei Dennis D Göke Jonathan J Tan Patrick P

Genome biology 20210122 1

<h4>Background</h4>Deregulated gene expression is a hallmark of cancer; however, most studies to date have analyzed short-read RNA sequencing data with inherent limitations. Here, we combine PacBio long-read isoform sequencing (Iso-Seq) and Illumina paired-end short-read RNA sequencing to comprehensively survey the transcriptome of gastric cancer (GC), a leading cause of global cancer mortality.<h4>Results</h4>We performed full-length transcriptome analysis across 10 GC cell lines covering four ...[more]

PMID: 33482911

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Project description:Genome-wide association studies (GWASs) have revealed thousands of associations in many complex traits and diseases. Previous studies suggest that a subset of associations are due to alterations in splicing; however, interpreting the effects of splicing on protein isoforms is hindered by limitations in defining full-length transcript isoforms using short-read RNA-seq data. Long-read RNA-seq represents a powerful approach to define and quantify transcript isoforms. In this study, we developed a novel approach that integrates information from GWAS, splicing QTL (sQTL), and PacBio long-read RNA-seq in a disease relevant model to infer the effects of sQTL on the ultimate protein isoform products they encode. Such information enables identification of genes potentially responsible for GWAS associations. As a proof-of-concept, we generated deep coverage (N=~22 million full-length reads) PacBio long-read RNAseq data on human fetal osteoblasts (hFOBs), a cell-line of relevance to the regulation of bone mineral density (BMD). We identified 68,326 protein-coding isoforms, including 17,375 (25%) which were novel. Next, we used Bayesian colocalization to identify 1,863 sQTLs from the Genotype-Tissue Expression (GTEx) project in 732 protein-coding genes which colocalized with BMD associations (H4PP > 0.75). A total of 836 junctions with colocalizing sQTLs in 459 (of the 732) genes were expressed in hFOB long-read RNA-seq data. With these data, we formulated hypotheses regarding the potential mechanism of action of each sQTL. For example, we identified 7 junctions with colocalizing sQTLs (maximum H4PP = 0.98-0.99) in TPM2 for splice junctions between two nearly mutually exclusive exons, and two different transcript termination sites, making it impossible to interpret without long-read RNA-seq data. siRNA mediated knockdown in hFOBs showed two TPM2 isoforms with opposing effects on mineralization. Our results suggest that splicing is a major mechanism underlying GWAS associations and long-read proteogenomics data is critical to precisely define the protein isoforms that are produced from splicing alterations.

Dataset Information

Long-read transcriptome sequencing reveals abundant promoter diversity in distinct molecular subtypes of gastric cancer

Publications

Long-read transcriptome sequencing reveals abundant promoter diversity in distinct molecular subtypes of gastric cancer.

Similar Datasets

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