Talk abstracts
Talk on Friday 01:15-01:30pm submitted by Anthony Caputo
T-sncR-seq allows for robust detection of transfer RNAs and other small noncoding RNAs at single cell resolution
Anthony Caputo (Case Western Reserve University School of Medicine, Department of Genetics and Genome Sciences, Cleveland, Ohio, United States), Ashleigh Schaffer (Oregon Health & Science University School of Medicine, Department of Medical and Molecular Genetics, Portland, Oregon, United States), Thomas J. Sweet (Case Western Reserve University School of Medicine, Department of Nutrition, Cleveland, Ohio, United States)
Abstract:
Over the past 10-15 years, single cell RNA sequencing (sc-RNA-seq) has proven to be an incredible valuable research tool, providing unprecedented insight into the cellular heterogeneity of the transcriptome. However, one major limitation of sc-RNA-seq is that it almost exclusively captures messenger RNA (mRNA), which only comprises 2-5% of the transcriptome. The other 95% is noncoding RNA (ncRNA), a diverse collection of molecules that are just as important as mRNA for regulating cellular processes but are mostly invisible to current sc-RNA-seq methods. Transfer RNAs (tRNAs) are a subset of ncRNAs that comprise 10-15% of the transcriptome. While originally viewed as passive adapters that simply bring amino acids to the translating ribosome, recent research has revealed that tRNAs are active regulators of gene expression, protein integrity, and cell stress responses. These findings were made possible by a wave of bulk RNA-seq pipelines tailored for tRNA detection. However, no existing technologies can robustly detect tRNAs at single cell resolution.
To bridge this gap, we developed tRNA and small noncoding RNA sequencing (T-sncR-seq) to robustly detect tRNAs and other sncRNAs at cellular resolution. We based our methodology on the widely used, high throughput 10X Genomics Chromium platform. We applied T-sncR-seq to human embryonic stem cells (hESCs) to benchmark our approach against previously validated tRNA sequencing methods. We then applied T-sncR-seq to both resting and stimulated CD4+ T cells isolated from healthy donors and found that hundreds of ncRNAs, both tRNAs and other sncRNAs, radically change expression upon T cell activation. We can also detect sequencing variation likely attributed to tRNA modifications at the level of single cells. Currently, we are integrating T-sncR-seq with mRNA sequencing to provide complete transcriptomic information at cellular resolution.
Keywords: transfer RNA (tRNA), small noncoding RNA (sncRNA), single cell RNA sequencing