RNA-Seq analyses of isoalted primary cardiac fibroblasts from wild type and Fam114a1-/- mouse hearts

Purpose: The biological and biochemical functions of FAM114A1 in mouse hearts or cardiac fibroblasts are entirely unknown. The goal of this study is to use the next generation sequencing (NGS) RNA-Seq followed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) methods to identify the downstream effector genes that are regulated by FAM114A1 in isolated primary cardiac fibroblasts from the hearts of wild type (WT; C57BL6/J) and Fam114a1-/- (global knockout; under C57BL6/J genetic background) mice at the mRNA level. Using an optimized data analysis workflow from the Genomic Resource Center at the University of Rochester Medical Center, we analyzed the RNA-Seq data of primary cardiac fibroblasts from P60 WT and Fam114a1-/- mice. We identifed differentially regulated genes (upregulated or downregulated) with statistical significance at the mRNA level (padj <0.05 for differential expressed genes). We found 28 downregulated genes involved in inflammation, hypertension, and extracelluar matrix receptor interaction. Furthermore, 20 genes were found to be upregulated and related to ribosome and translation pathways. About half the downregulated gene were confirmed by qRT-PCR. This study is the first transcriptomic analysis of isolated primary cardiac fibroblasts of Fam114a1-/- versus WT mice with biological triplicates using RNA-Seq technology. The optimized data analysis workflows offer a framework for comparative investigations of expression profiles. Our RNA-Seq analysis and follow-up qRT-PCR validation results suggest that among all the downregualted genes ADAMTS15 is found to be a critical downstream effector gene of FAM114A1 and is important for cardiac fibrobalt activation and proliferation. Overall design: RNA-Seq of WT vs Fam114a1-/- mouse hearts