Eschenbach, J. | Search bioRxiv

bioRxiv2024

Transverse Aortic COnstriction Multi-omics Analysis (TACOMA) uncovers pathophysiological cardiac molecular mechanisms

Time-course multi-omics data of a murine model of progressive heart failure induced by transverse aortic constriction (TAC) provide insights into the molecular mechanisms that are causatively involved in contractile failure and structural cardiac remodelling. We employ Illumina-based transcriptomics, Nanopore sequencing, and mass spectrometry-based proteomics on samples from the left ventricle (LV) and right ventricle (RV, RNA only) of the heart at 1, 7, 21, and 56 days following TAC and Sham surgery. Here, we present TACOMA, as an interactive web-application that integrates and visualizes transcriptomics and proteomics data collected in a TAC time-course experiment. TACOMA enables users to visualize the expression profile of known and novel genes and protein products thereof. Importantly, we capture alternative splicing events by assessing differential transcript and exon usage as well. Co-expression-based clustering algorithms and functional enrichment analysis revealed overrepresented annotations of biological processes and molecular functions at the protein and gene levels. To enhance data integration, TACOMA synchronizes transcriptomics and proteomics profiles, enabling cross-omics comparisons. With TACOMA (https://shiny.dieterichlab.org/app/tacoma), we offer a rich web-based resource to uncover molecular events and biological processes implicated in contractile failure and cardiac hypertrophy. For example, we highlight: (i) changes in metabolic genes and proteins in the time course of hypertrophic growth and contractile impairment; (ii) identification of RNA splicing changes in the expression of Tpm2 isoforms between RV and LV; and (iii) novel transcripts and genes likely contributing to the pathogenesis of heart failure. We plan to extend these data with additional environmental and genetic models of heart failure to decipher common and distinct molecular changes in heart diseases of different aetiologies. Database URLhttps://shiny.dieterichlab.org/app/tacoma

bioinformatics↗

bioRxiv2021

Deep assessment of human disease-associated ribosomal RNA modifications using Nanopore direct RNA sequencing

The catalytically active component of ribosomes, rRNA, is long studied and heavily modified. However, little is known about functional and pathological consequences of changes in human rRNA modification status. Direct RNA sequencing on the Nanopore platform enables the direct assessment of rRNA modifications. We established a targeted Nanopore direct rRNA sequencing approach and applied it to CRISPR-Cas9 engineered HCT116 cells, lacking specific enzymatic activities required to establish defined rRNA base modifications. We analyzed these sequencing data along with wild type samples and in vitro transcribed reference sequences to specifically detect changes in modification status. We show for the first time that direct RNA-sequencing is feasible on smaller, i.e. Flongle, flow cells. Our targeted approach reduces RNA input requirements, making it accessible to the analysis of limited samples such as patient derived material. The analysis of rRNA modifications during cardiomyocyte differentiation of human induced pluripotent stem cells, and of heart biopsies from cardiomyopathy patients revealed altered modifications of specific sites, among them pseudouridines, 2-O-methylation of riboses and acetylation of cytidines. Targeted direct rRNA-seq analysis with JACUSA2 opens up the possibility to analyze dynamic changes in rRNA modifications in a wide range of biological and clinical samples.

molecular biology↗