ABSTRACT
The heterogeneous composition of cellular transcriptomes poses a major challenge for detecting weakly expressed RNA classes, as they can be obscured by abundant RNAs. Although biochemical protocols can enrich or deplete specified RNAs, they are time-consuming, expensive and can compromise RNA integrity. Here we introduce RISER, a biochemical-free technology for the real-time enrichment or depletion of RNA classes. RISER performs selective rejection of molecules during direct RNA sequencing by identifying RNA classes directly from nanopore signals with deep learning and communicating with the sequencing hardware in real time. By targeting the dominant messenger and mitochondrial RNA classes for depletion, RISER reduces their respective read counts by more than 85%, resulting in an increase in sequencing depth of 47% on average for long non-coding RNAs. We also apply RISER for the depletion of globin mRNA in whole blood, achieving a decrease in globin reads by more than 90% as well as an increase in non-globin reads by 16% on average. Furthermore, using a GPU or a CPU, RISER is faster than GPU-accelerated basecalling and mapping. RISER's modular and retrainable software and intuitive command-line interface allow easy adaptation to other RNA classes. RISER is available at https://github.com/comprna/riser .
Subject(s)
RNA, Messenger , Sequence Analysis, RNA , Sequence Analysis, RNA/methods , Humans , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA, Long Noncoding/genetics , RNA/genetics , Software , Globins/genetics , High-Throughput Nucleotide Sequencing/methods , Deep Learning , Transcriptome , RNA, Mitochondrial/genetics , RNA, Mitochondrial/metabolismABSTRACT
The phosphoprotein AHNAK is a large, ubiquitously expressed scaffolding protein involved in mediating a host of protein-protein interactions. This enables AHNAK to participate in various multi-protein complexes thereby orchestrating a range of diverse biological processes, including tumour suppression, immune regulation and cell architecture maintenance. A less studied but nonetheless equally important function occurs in calcium homeostasis. It does so by largely interacting with the L-type voltage-gated calcium channel (LVGCC) present in the plasma membrane of excitable cells such as muscles and neurons. Several studies have characterized the underlying basis of AHNAK's functional role in calcium channel modulation, which has led to a greater understanding of this cellular process and its associated pathologies. In this article we review and examine recent advances relating to the physiological aspects of AHNAK in calcium regulation. Specifically, we will provide a broad overview of AHNAK including its structural makeup and its interaction with several isoforms of LVGCC, and how these molecular interactions regulate calcium modulation across various tissues and their implication in muscle and neuronal function.