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With increased usage of long-read sequencing technologies to perform transcriptome analyses, there becomes a greater need to evaluate different methodologies including library preparation, sequencing platform, and computational analysis tools. Here, we report the study design of a community effort called the Long-read RNA-Seq Genome Annotation Assessment Project (LRGASP) Consortium, whose goals are characterizing the strengths and remaining challenges in using long-read approaches to identify and quantify the transcriptomes of both model and non-model organisms. The LRGASP organizers have generated cDNA and direct RNA datasets in human, mouse, and manatee samples using different protocols followed by sequencing on Illumina, Pacific Biosciences, and Oxford Nanopore Technologies platforms. Participants will use the provided data to submit predictions for three challenges: transcript isoform detection with a high-quality genome, transcript isoform quantification, and de novo transcript isoform identification. Evaluators from different institutions will determine which pipelines have the highest accuracy for a variety of metrics using benchmarks that include spike-in synthetic transcripts, simulated data, and a set of undisclosed, manually curated transcripts by GENCODE. We also describe plans for experimental validation of predictions that are platform-specific and computational tool-specific. We believe that a community effort to evaluate long-read RNA-seq methods will help move the field toward a better consensus on the best approaches to use for transcriptome analyses.

Pore forming toxins (PFTs) are virulent proteins released by several species, including many strains of bacteria, to attack and kill host cells. In this article, we focus on the utility of molecular dynamics (MD) simulations and the molecular insights gleaned from these techniques on the pore forming pathways of PFTs. In addition to all-atom simulations which are widely used, coarse-grained MARTINI models and structure-based models have also been used to study PFTs. Here, the emphasis is on methods and techniques involved while setting up, monitoring, and evaluating properties from MD simulations of PFTs in a membrane environment. We draw from several case studies to illustrate how MD simulations have provided molecular insights into protein-protein and protein-lipid interactions, lipid dynamics, conformational transitions and structures of both the oligomeric intermediates and assembled pore structures.

RAS genes are the most frequently mutated oncogenes in cancer, yet the effects of oncogenic RAS signaling on the noncoding transcriptome remain unclear. We analyzed the transcriptomes of human airway and bronchial epithelial cells transformed with mutant KRAS to define the landscape of KRAS-regulated noncoding RNAs. We find that oncogenic KRAS signaling upregulates noncoding transcripts throughout the genome, many of which arise from transposable elements (TEs). These TE RNAs exhibit differential expression, are preferentially released in extracellular vesicles, and are regulated by KRAB zinc-finger (KZNF) genes, which are broadly downregulated in mutant KRAS cells and lung adenocarcinomas in vivo. Moreover, mutant KRAS induces an intrinsic IFN-stimulated gene (ISG) signature that is often seen across many different cancers. Our results indicate that mutant KRAS remodels the repetitive noncoding transcriptome, demonstrating the broad scope of intracellular and extracellular RNAs regulated by this oncogenic signaling pathway.

Pore-forming toxins (PFTs) form nanoscale pores across target membranes causing cell death. Cytolysin A (ClyA) from Escherichia coli is a prototypical alpha-helical toxin that contributes to cytolytic phenotype of several pathogenic strains. It is produced as a monomer and, upon membrane exposure, undergoes conformational changes and finally oligomerizes to form a dodecameric pore, thereby causing ion imbalance and finally cell death. However, our current understanding of this assembly process is limited to studies in detergents, which do not capture the physicochemical properties of biological membranes. Here, using single-molecule imaging and molecular dynamics simulations, we study the ClyA assembly pathway on phospholipid bilayers. We report that cholesterol stimulates pore formation, not by enhancing initial ClyA binding to the membrane but by selectively stabilizing a protomer-like conformation. This was mediated by specific interactions by cholesterol-interacting residues in the N-terminal helix. Additionally, cholesterol stabilized the oligomeric structure using bridging interactions in the protomer-protomer interfaces, thereby resulting in enhanced ClyA oligomerization. This dual stabilization of distinct intermediates by cholesterol suggests a possible molecular mechanism by which ClyA achieves selective membrane rupture of eukaryotic cell membranes. Topological similarity to eukaryotic membrane proteins suggests evolution of a bacterial alpha-toxin to adopt eukaryotic motifs for its activation. Broad mechanistic correspondence between pore-forming toxins hints at a wider prevalence of similar protein membrane insertion mechanisms.

In 2013, Farid and Vasiliev [ arXiv:1310.4922 [quant-ph]] for the first time proposed a way to construct a protocol for the realisation of “Classical to Quantum” one-way hash function, a derivative of the quantum one-way function as defined by Gottesman and Chuang [Technical Report arXiv:quant-ph/0105032 ] and used it for constructing quantum digital signatures. We, on the other hand, for the first time, propose the idea of a different kind of one-way function, which is “quantum-classical” in nature, that is, it takes an n-qubit quantum state of a definite kind as its input and produces a classical output. We formally define such a one-way function and propose a way to construct and realise it. The proposed one-way function turns out to be very useful in authenticating a quantum state in any quantum money scheme, and so we can construct many different quantum money schemes based on such a one-way function. Later in the paper, we also give explicit constructions of some interesting quantum money schemes like quantum bitcoins and quantum currency schemes, solely based on the proposed one-way function. The security of such schemes can be explained on the basis of the security of the underlying one-way functions.