Abolished frameshifting for predicted structure-stabilizing SARS-CoV-2 mutants: implications to alternative conformations and their statistical structural analyses.

The SARS-CoV-2 frameshifting element (FSE) has been intensely studied and explored as a therapeutic target for coronavirus diseases, including COVID-19. Besides the intriguing virology, this small RNA is known to adopt many length-dependent conformations, as verified by multiple experimental and computational approaches. However, the role these alternative conformations play in the frameshifting mechanism and how to quantify this structural abundance has been an ongoing challenge.

Visualizing RNA structure ensembles by single-molecule correlated chemical probing.

RNA molecules fold to form complex internal structures. Many of these RNA structures populate ensembles with rheostat-like properties, with each state having a distinct function. Until recently, analysis of RNA structures, especially within cells, was limited to modeling either a single averaged structure or computationally-modeled ensembles.

An intricate balancing act: Upstream and downstream frameshift co-regulatory elements.

Targeting ribosomal frameshifting has emerged as a potential therapeutic intervention strategy against Covid-19. During ribosomal translation, a fraction of elongating ribosomes slips by one base in the 5' direction and enters a new reading frame for viral protein synthesis. Any interference with this process profoundly affects viral replication and propagation.

Orb2 enables rare-codon-enriched mRNA expression during Drosophila neuron differentiation.

Regulation of codon optimality is an increasingly appreciated layer of cell- and tissue-specific protein expression control. Here, we use codon-modified reporters to show that differentiation of Drosophila neural stem cells into neurons enables protein expression from rare-codon-enriched genes. From a candidate screen, we identify the cytoplasmic polyadenylation element binding (CPEB) protein Orb2 as a positive regulator of rare-codon-dependent mRNA stability in neurons.

Abolished frameshifting for predicted structure-stabilizing SARS-CoV-2 mutants: Implications to alternative conformations and their statistical structural analyses.

The SARS-CoV-2 frameshifting element (FSE) has been intensely studied and explored as a therapeutic target for coronavirus diseases including COVID-19. Besides the intriguing virology, this small RNA is known to adopt many length-dependent conformations, as verified by multiple experimental and computational approaches. However, the role these alternative conformations play in the frameshifting mechanism and how to quantify this structural abundance has been an ongoing challenge.

Colocalization analysis of 3' UTR alternative polyadenylation quantitative trait loci reveals novel mechanisms underlying associations with lung function.

While many disease-associated single nucleotide polymorphisms (SNPs) are expression quantitative trait loci (eQTLs), a large proportion of genome-wide association study (GWAS) variants are of unknown function. Alternative polyadenylation (APA) plays an important role in posttranscriptional regulation by allowing genes to shorten or extend 3' untranslated regions (UTRs). We hypothesized that genetic variants that affect APA in lung tissue may lend insight into the function of respiratory associated GWAS loci.

Coordinated expression of replication-dependent histone genes from multiple loci promotes histone homeostasis in .

Production of large amounts of histone proteins during S phase is critical for proper chromatin formation and genome integrity. This process is achieved in part by the presence of multiple copies of replication dependent (RD) histone genes that occur in one or more clusters in metazoan genomes. In addition, RD histone gene clusters are associated with a specialized nuclear body, the histone locus body (HLB), which facilitates efficient transcription and 3' end-processing of RD histone mRNA.

Orb2 enables rare-codon-enriched mRNA expression during neuron differentiation.

Regulation of codon optimality is an increasingly appreciated layer of cell- and tissue-specific protein expression control. Here, we use codon-modified reporters to show that differentiation of neural stem cells into neurons enables protein expression from rare-codon-enriched genes. From a candidate screen, we identify the cytoplasmic polyadenylation element binding (CPEB) protein Orb2 as a positive regulator of rare-codon-dependent expression in neurons.

Structural analysis of MALAT1 long noncoding RNA in cells and in evolution.

Although not canonically polyadenylated, the long noncoding RNA MALAT1 (metastasis-associated lung adenocarcinoma transcript 1) is stabilized by a highly conserved 76-nt triple helix structure on its 3' end. The entire MALAT1 transcript is over 8000 nt long in humans. The strongest structural conservation signal in MALAT1 (as measured by covariation of base pairs) is in the triple helix structure.

Global 5'-UTR RNA structure regulates translation of a SERPINA1 mRNA.

SERPINA1 mRNAs encode the protease inhibitor α-1-antitrypsin and are regulated through post-transcriptional mechanisms. α-1-antitrypsin deficiency leads to chronic obstructive pulmonary disease (COPD) and liver cirrhosis, and specific variants in the 5'-untranslated region (5'-UTR) are associated with COPD. The NM_000295.

Rotavirus RNA chaperone mediates global transcriptome-wide increase in RNA backbone flexibility.

Due to genome segmentation, rotaviruses must co-package eleven distinct genomic RNAs. The packaging is mediated by virus-encoded RNA chaperones, such as the rotavirus NSP2 protein. While the activities of distinct RNA chaperones are well studied on smaller RNAs, little is known about their global effect on the entire viral transcriptome.

Quantitative prediction of variant effects on alternative splicing in using endogenous pre-messenger RNA structure probing.

Splicing is highly regulated and is modulated by numerous factors. Quantitative predictions for how a mutation will affect precursor mRNA (pre-mRNA) structure and downstream function are particularly challenging. Here, we use a novel chemical probing strategy to visualize endogenous precursor and mature mRNA structures in cells.

Distinct responses to rare codons in select tissues.

Codon usage bias has long been appreciated to influence protein production. Yet, relatively few studies have analyzed the impacts of codon usage on tissue-specific mRNA and protein expression. Here, we use codon-modified reporters to perform an organism-wide screen in for distinct tissue responses to codon usage bias.

Sequence and tissue targeting specificity of ZFP36L2 reveals Elavl2 as a novel target with co-regulation potential.

Zinc finger protein 36 like 2 (ZFP36L2) is an RNA-binding protein that destabilizes transcripts containing adenine-uridine rich elements (AREs). The overlap between ZFP36L2 targets in different tissues is minimal, suggesting that ZFP36L2-targeting is highly tissue specific. We developed a novel Zfp36l2-lacking mouse model (L2-fKO) to identify factors governing this tissue specificity.

Identifying proximal RNA interactions from cDNA-encoded crosslinks with ShapeJumper.

SHAPE-JuMP is a concise strategy for identifying close-in-space interactions in RNA molecules. Nucleotides in close three-dimensional proximity are crosslinked with a bi-reactive reagent that covalently links the 2'-hydroxyl groups of the ribose moieties. The identities of crosslinked nucleotides are determined using an engineered reverse transcriptase that jumps across crosslinked sites, resulting in a deletion in the cDNA that is detected using massively parallel sequencing.

A novel algorithm for ranking RNA structure candidates.

RNA research is advancing at an ever increasing pace. The newest and most state-of-the-art instruments and techniques have made possible the discoveries of new RNAs, and they have carried the field to new frontiers of disease research, vaccine development, therapeutics, and architectonics. Like proteins, RNAs show a marked relationship between structure and function.

In vivo architecture of the telomerase RNA catalytic core in Trypanosoma brucei.

Telomerase is a unique ribonucleoprotein (RNP) reverse transcriptase that utilizes its cognate RNA molecule as a template for telomere DNA repeat synthesis. Telomerase contains the reverse transcriptase protein, TERT and the template RNA, TR, as its core components. The 5'-half of TR forms a highly conserved catalytic core comprising of the template region and adjacent domains necessary for telomere synthesis.

Alternative poly-adenylation modulates α1-antitrypsin expression in chronic obstructive pulmonary disease.

α1-anti-trypsin (A1AT), encoded by SERPINA1, is a neutrophil elastase inhibitor that controls the inflammatory response in the lung. Severe A1AT deficiency increases risk for Chronic Obstructive Pulmonary Disease (COPD), however, the role of A1AT in COPD in non-deficient individuals is not well known. We identify a 2.

Cigarette smoking-associated isoform switching and 3' UTR lengthening via alternative polyadenylation.

Cigarette smoking induces a profound transcriptomic and systemic inflammatory response. Previous studies have focused on gene level differential expression of smoking, but the genome-wide effects of smoking on alternative isoform regulation have not yet been described. We conducted RNA sequencing in whole-blood samples of 454 current and 767 former smokers in the COPDGene Study, and we analyzed the effects of smoking on differential usage of isoforms and exons.

Disease-associated human genetic variation through the lens of precursor and mature RNA structure.

Disease-associated variants (DAVs) are commonly considered either through a genomic lens that describes variant function at the DNA level, or at the protein function level if the variant is translated. Although the genomic and proteomic effects of variation are well-characterized, genetic variants disrupting post-transcriptional regulation is another mechanism of disease that remains understudied. Specific RNA sequence motifs mediate post-transcriptional regulation both in the nucleus and cytoplasm of eukaryotic cells, often by binding to RNA-binding proteins or other RNAs.

To Knot or Not to Knot: Multiple Conformations of the SARS-CoV-2 Frameshifting RNA Element.

The SARS-CoV-2 frameshifting RNA element (FSE) is an excellent target for therapeutic intervention against Covid-19. This small gene element employs a shifting mechanism to pause and backtrack the ribosome during translation between Open Reading Frames 1a and 1b, which code for viral polyproteins. Any interference with this process has a profound effect on viral replication and propagation.

Direct Mapping of Higher-Order RNA Interactions by SHAPE-JuMP.

Higher-order structure governs function for many RNAs. However, discerning this structure for large RNA molecules in solution is an unresolved challenge. Here, we present SHAPE-JuMP (selective 2'-hydroxyl acylation analyzed by primer extension and juxtaposed merged pairs) to interrogate through-space RNA tertiary interactions.

To knot or not to knot: Multiple conformations of the SARS-CoV-2 frameshifting RNA element.

The SARS-CoV-2 frameshifting RNA element (FSE) is an excellent target for therapeutic intervention against Covid-19. This small gene element employs a shifting mechanism to pause and backtrack the ribosome during translation between Open Reading Frames 1a and 1b, which code for viral polyproteins. Any interference with this process has profound effect on viral replication and propagation.