Biology subjects

Lee, Y.-k.

Publications and source records attributed to Lee, Y.-k..

3 recordsLinked to original sources

bioRxiv2024

Cellular dsRNA interactome reveals the regulatory map of exogenous RNA sensing

RNA-binding proteins (RBPs) provide a critical post-transcriptional regulatory layer in determining RNA fate. Currently, UV crosslinking followed by oligo-dT pull-down is the golden standard in identifying the RBP repertoire, but such method is ineffective in capturing RBPs that recognize double-stranded RNAs (dsRNAs). Here, we utilize anti-dsRNA antibody immunoprecipitation followed by quantitative mass spectrometry to comprehensively identify RBPs bound to cellular dsRNAs without any external stimulus. Notably, our dsRNA interactome contains proteins involved in sensing N6-methyladenosine RNAs and stress granule components. We further perform targeted CRISPR-Cas9 knockout functional screening and discover proteins that can regulate the interferon (IFN) response during exogenous RNA sensing. Interestingly, depleting dsRBPs mostly attenuate the IFN response, and they act as antiviral factors during human {beta}-coronavirus HCoV-OC43 infection. Our dsRNA interactome capture provides an unbiased and comprehensive characterization of putative dsRBPs and will facilitate our understanding of dsRNA sensing in both physiological and pathological contexts.

molecular biology↗

bioRxiv2024

SLIRP mediates nuclear-mitochondrial communication during antiviral signaling to promote robust interferon response

The abnormal innate immune response is a prominent feature underlying autoimmune diseases. One emerging factor that can trigger dysregulated immune activation is cytosolic mitochondrial double-stranded RNAs (mt-dsRNAs). However, the mechanism by which mt-dsRNAs stimulate immune responses remains poorly understood. Here, we discover SRA stem-loop interacting RNA binding protein (SLIRP) as a key amplifier of mt-dsRNA-triggered antiviral signals. In autoimmune diseases, SLIRP is commonly upregulated, and targeted knockdown of SLIRP dampens the interferon response. We find that the activation of melanoma differentiation-associated gene 5 (MDA5) by exogenous dsRNAs upregulates SLIRP, which then stabilizes mt-dsRNAs and promotes their cytosolic release to activate MDA5 further, augmenting the interferon response. Furthermore, the downregulation of SLIRP partially rescues the abnormal interferon-stimulated gene expression in autoimmune patients primary cells and makes cells vulnerable to certain viral infections. Our study unveils SLIRP as a pivotal mediator of interferon response through positive feedback amplification of antiviral signaling.

molecular biology↗

bioRxiv2024

Revealing nanoscale structure and interfaces of protein and polymer condensates via cryo-electron microscopy

Liquid-liquid phase separation (LLPS) is a ubiquitous demixing phenomenon observed in various molecular solutions, including in polymer and protein solutions. Demixing of solutions results in condensed, phase separated droplets which exhibit a range of liquid-like properties driven by transient intermolecular interactions. Understanding the organization within these condensates is crucial for deciphering their material properties and functions. This study explores the distinct nanoscale networks and interfaces in the condensate samples using a modified cryo-electron microscopy (cryo-EM) method. The method involves initiating condensate formation on electron microscopy grids to control droplet size and stage in the phase separation process. The versatility of this method is demonstrated by imaging three different classes of condensates. We further investigate the condensate structures using cryo-electron tomography which provides 3D reconstructions, uncovering porous internal structures, unique core-shell morphologies, and inhomogeneities within the nanoscale organization of protein condensates. Comparison with dry-state transmission electron microscopy emphasizes the importance of preserving the hydrated structure of condensates for accurate structural analysis. We correlate the internal structure of protein condensates with their amino acid sequences and material properties by performing viscosity measurements that support that more viscous condensates exhibit denser internal assemblies. Our findings contribute to a comprehensive understanding of nanoscale condensate structure and its material properties. Our approach here provides a versatile tool for exploring various phase-separated systems and their nanoscale structures for future studies.

biophysics↗