Protocol for profiling virus-to-host RNA-RNA interactions in infected cells by RIC-seq.

Virus-to-host RNA-RNA interactions directly regulate host mRNA stability and viral replication. However, globally profiling virus-to-host in situ RNA-RNA interactions remains challenging. Here, we present an RNA in situ conformation sequencing (RIC-seq)-based protocol for mapping high-confidence virus-to-host in situ RNA-RNA interactions in infected cells.

SARS-CoV-2 RNA stabilizes host mRNAs to elicit immunopathogenesis.

SARS-CoV-2 RNA interacts with host factors to suppress interferon responses and simultaneously induces cytokine release to drive the development of severe coronavirus disease 2019 (COVID-19). However, how SARS-CoV-2 hijacks host RNAs to elicit such imbalanced immune responses remains elusive. Here, we analyzed SARS-CoV-2 RNA in situ structures and interactions in infected cells and patient lung samples using RIC-seq.

Complementary Alu sequences mediate enhancer-promoter selectivity.

Enhancers determine spatiotemporal gene expression programs by engaging with long-range promoters. However, it remains unknown how enhancers find their cognate promoters. We recently developed a RNA in situ conformation sequencing technology to identify enhancer-promoter connectivity using pairwise interacting enhancer RNAs and promoter-derived noncoding RNAs.

Structural insights reveal the specific recognition of meiRNA by the Mei2 protein.

In the fission yeast Schizosaccharomyces pombe, Mei2, an RNA-binding protein essential for entry into meiosis, regulates meiosis initiation. Mei2 binds to a specific non-coding RNA species, meiRNA, and accumulates at the sme2 gene locus, which encodes meiRNA. Previous research has shown that the Mei2 C-terminal RNA recognition motif (RRM3) physically interacts with the meiRNA 5' region in vitro and stimulates meiosis in vivo.

Emerging roles of RNA-RNA interactions in transcriptional regulation.

Pervasive transcription of the human genome generates a massive amount of noncoding RNAs (ncRNAs) that lack protein-coding potential but play crucial roles in development, differentiation, and tumorigenesis. To achieve these biological functions, ncRNAs must first fold into intricate structures via intramolecular RNA-RNA interactions (RRIs) and then interact with different RNA substrates via intermolecular RRIs. RRIs are usually facilitated, stabilized, or mediated by RNA-binding proteins.

The architecture of the SARS-CoV-2 RNA genome inside virion.

SARS-CoV-2 carries the largest single-stranded RNA genome and is the causal pathogen of the ongoing COVID-19 pandemic. How the SARS-CoV-2 RNA genome is folded in the virion remains unknown. To fill the knowledge gap and facilitate structure-based drug development, we develop a virion RNA in situ conformation sequencing technology, named vRIC-seq, for probing viral RNA genome structure unbiasedly.

Global profiling of RNA-binding protein target sites by LACE-seq.

RNA-binding proteins (RBPs) have essential functions during germline and early embryo development. However, current methods are unable to identify the in vivo targets of a RBP in these low-abundance cells. Here, by coupling RBP-mediated reverse transcription termination with linear amplification of complementary DNA ends and sequencing, we present the LACE-seq method for identifying RBP-regulated RNA networks at or near the single-oocyte level.

Global in situ profiling of RNA-RNA spatial interactions with RIC-seq.

Emerging evidence has demonstrated that RNA-RNA interactions are vital in controlling diverse biological processes, including transcription, RNA splicing and protein translation. RNA in situ conformation sequencing (RIC-seq) is a technique for capturing protein-mediated RNA-RNA proximal interactions globally in living cells at single-base resolution. Cells are first treated with formaldehyde to fix all the protein-mediated RNA-RNA interactions in situ.

RIC-seq for global in situ profiling of RNA-RNA spatial interactions.

Highly structured RNA molecules usually interact with each other, and associate with various RNA-binding proteins, to regulate critical biological processes. However, RNA structures and interactions in intact cells remain largely unknown. Here, by coupling proximity ligation mediated by RNA-binding proteins with deep sequencing, we report an RNA in situ conformation sequencing (RIC-seq) technology for the global profiling of intra- and intermolecular RNA-RNA interactions.

RNA-binding protein DDX1 is responsible for fatty acid-mediated repression of insulin translation.

The molecular mechanism in pancreatic β cells underlying hyperlipidemia and insulin insufficiency remains unclear. Here, we find that the fatty acid-induced decrease in insulin levels occurs due to a decrease in insulin translation. Since regulation at the translational level is generally mediated through RNA-binding proteins, using RNA antisense purification coupled with mass spectrometry, we identify a novel insulin mRNA-binding protein, namely, DDX1, that is sensitive to palmitate treatment.

The RNA-binding protein ROD1/PTBP3 cotranscriptionally defines AID-loading sites to mediate antibody class switch in mammalian genomes.

Activation-induced cytidine deaminase (AID) mediates class switching by binding to a small fraction of single-stranded DNA (ssDNA) to diversify the antibody repertoire. The precise mechanism for highly selective AID targeting in the genome has remained elusive. Here, we report an RNA-binding protein, ROD1 (also known as PTBP3), that is both required and sufficient to define AID-binding sites genome-wide in activated B cells.

Juvenile hormone-dependent Kazal-type serine protease inhibitor Greglin safeguards insect vitellogenesis and egg production.

In addition to preventing insect metamorphosis, juvenile hormone (JH) is known to stimulate aspects of insect reproduction. However, the molecular mechanisms of JH action in insect reproduction remain largely unknown. By reanalyzing the transcriptomic data from adults and other developmental stages of the migratory locust Locusta migratoria, we identified a gene coding for Kazal-type protease inhibitor, previously named Greglin.

Arabidopsis ARGONAUTE 1 Binds Chromatin to Promote Gene Transcription in Response to Hormones and Stresses.

Conventional RNA interference (RNAi) pathways suppress eukaryotic gene expression at the transcriptional or post-transcriptional level. At the core of RNAi are small RNAs (sRNAs) and effector Argonaute (AGO) proteins. Arabidopsis AGO1 is known to bind microRNAs (miRNAs) and post-transcriptionally repress target genes in the cytoplasm.