Prediction of Metabolic Profiles from Transcriptomics Data in Human Cancer Cell Lines.

The Metabolome and Transcriptome are mutually communicating within cancer cells, and this interplay is translated into the existence of quantifiable correlation structures between gene expression and metabolite abundance levels. Studying these correlations could provide a novel venue of understanding cancer and the discovery of novel biomarkers and pharmacological strategies, as well as laying the foundation for the prediction of metabolite quantities by leveraging information from the more widespread transcriptomics data. In the current paper, we investigate the correlation between gene expression and metabolite levels in the Cancer Cell Line Encyclopedia dataset, building a direct correlation network between the two molecular ensembles. We show that a metabolite/transcript correlation network can be used to predict metabolite levels in different samples and datasets, such as the NCI-60 cancer cell line dataset, both on a sample-by-sample basis and in differential contrasts. We also show that metabolite levels can be predicted in principle on any sample and dataset for which transcriptomics data are available, such as the Cancer Genome Atlas (TCGA).

The NGS WikiBook: a dynamic collaborative online training effort with long-term sustainability.

Next-generation sequencing (NGS) is increasingly being adopted as the backbone of biomedical research. With the commercialization of various affordable desktop sequencers, NGS will be reached by increasing numbers of cellular and molecular biologists, necessitating community consensus on bioinformatics protocols to tackle the exponential increase in quantity of sequence data. The current resources for NGS informatics are extremely fragmented. Finding a centralized synthesis is difficult. A multitude of tools exist for NGS data analysis; however, none of these satisfies all possible uses and needs. This gap in functionality could be filled by integrating different methods in customized pipelines, an approach helped by the open-source nature of many NGS programmes. Drawing from community spirit and with the use of the Wikipedia framework, we have initiated a collaborative NGS resource: The NGS WikiBook. We have collected a sufficient amount of text to incentivize a broader community to contribute to it. Users can search, browse, edit and create new content, so as to facilitate self-learning and feedback to the community. The overall structure and style for this dynamic material is designed for the bench biologists and non-bioinformaticians. The flexibility of online material allows the readers to ignore details in a first read, yet have immediate access to the information they need. Each chapter comes with practical exercises so readers may familiarize themselves with each step. The NGS WikiBook aims to create a collective laboratory book and protocol that explains the key concepts and describes best practices in this fast-evolving field.

Quantitative and qualitative detection of tRNAs, tRNA halves and tRFs in human cancer samples: Molecular grounds for biomarker development and clinical perspectives.

Transfer RNAs (tRNAs) are small non-coding RNAs playing a central role during protein synthesis. Besides translation, growing evidence suggests that in many contexts, precursor or mature tRNAs can also be processed into smaller fragments playing many non-canonical regulatory roles in different biological pathways with oncogenic relevance. Depending on the source, these molecules can be classified as tRNA halves (also known as tiRNAs) or tRNA-derived fragments (tRFs), and furtherly divided into 5'-tRNA and 3'-tRNA halves, or tRF-1, tRF-2, tRF-3, tRF-5, and i-tRF, respectively. Unlike DNA and mRNA, high-throughput sequencing of tRNAs is challenging, because of technical limitations of currently developed sequencing methods. In recent years, different sequencing approaches have been proposed allowing the quantification and identification of an increasing number of tRNA fragments with critical functions in distinct physiological and pathophysiological processes. In the present review, we discussed pros and cons of recent advances in different sequencing methods, also introducing the expanding repertoire of bioinformatics tool and resources specifically focused on tRNA research and discussing current issues in the study of these small RNA molecules. Furthermore, we discussed the potential value of tRNA fragments as diagnostic and prognostic biomarkers for different types of cancers.

Transcriptional Patterns in Stages of Alzheimer's Disease are Cell Type Specific and Partially Converge with the Effects of Alcohol Use Disorder in Humans.

Advances in single cell technologies have led to the discovery and characterization of new brain cell-types, that in turn lead to a better understanding of the pathogenesis of Alzheimer's disease (AD). Here, we present a detailed analysis of single nucleus (sn)RNA-seq data for three stages of AD from middle temporal gyrus (MTG) and compare it with snRNA-seq data from the prefrontal cortices from individuals with alcohol use disorder (AUD). We observed a significant decrease in both inhibitory and excitatory neurons, in general agreement with previous reports. We observed several cell-type specific gene expressions and pathways dysregulations that delineate AD stages. Endothelial and VLMCs (vascular leptomeningeal cells) showed the greatest degree of gene expression changes. Cell-type specific evidence of neurodegeneration was seen in multiple neuronal cell-types particularly in SST (somatostatin) and L5 ET (layer-5 extra-telencephalic) neurons, among others. Evidence of inflammatory responses were seen in non-neuronal cells, particularly in intermediate and advanced AD. We observed common perturbations in AD and AUD, particularly in pathways, like transcription, translation, apoptosis, autophagy, calcium signaling, neuroinflammation, and phosphorylation, that imply shared transcriptional pathogenic mechanisms and support a role for excessive alcohol intake in AD progression. Major AUD gene markers form and perturb a network of genes significantly associated with intermediate and advanced AD. Master regulator analysis from AUD gene markers revealed significant correlation with advanced AD of transcription factors that have implications in intellectual disability, neuroinflammation, and other neurodegenerative conditions, further suggesting a shared nexus of transcriptional changes between AD and AUD.Significance Statement This study holds significant implications for understanding the intricate molecular landscape of Alzheimer's disease (AD) and its intersection with alcohol use disorder (AUD). By profiling transcriptional changes in the neocortex associated with AD progression and comparing them with those in AUD, we shed light on shared gene expression and pathway dysregulations between the two conditions. Our findings corroborate prior research on neuronal depletion and highlight novel insights into cell-type-specific gene expression patterns in AD stages. Moreover, the identification of common genetic signatures suggests a potential exacerbating effect of AUD on AD progression. This comprehensive analysis not only deepens our understanding of AD pathology but also underscores the importance of considering AUD as a potential risk factor for accelerating AD onset or severity.

Cross-Species Transcriptomics Analysis Highlights Conserved Molecular Responses to Per- and Polyfluoroalkyl Substances.

In recent decades, per- and polyfluoroalkyl substances (PFASs) have garnered widespread public attention due to their persistence in the environment and detrimental effects on the health of living organisms, spurring the generation of several transcriptome-centered investigations to understand the biological basis of their mechanism. In this study, we collected 2144 publicly available samples from seven distinct animal species to examine the molecular responses to PFAS exposure and to determine if there are conserved responses. Our comparative transcriptional analysis revealed that exposure to PFAS is conserved across different tissues, molecules and species. We identified and reported several genes exhibiting consistent and evolutionarily conserved transcriptional response to PFASs, such as ESR1, HADHA and ID1, as well as several pathways including lipid metabolism, immune response and hormone pathways. This study provides the first evidence that distinct PFAS molecules induce comparable transcriptional changes and affect the same metabolic processes across inter-species borders. Our findings have significant implications for understanding the impact of PFAS exposure on living organisms and the environment. We believe that this study offers a novel perspective on the molecular responses to PFAS exposure and provides a foundation for future research into developing strategies for mitigating the detrimental effects of these substances in the ecosystem.

Structural genetics of circulating variants affecting the SARS-CoV-2 spike/human ACE2 complex.

SARS-CoV-2 entry in human cells is mediated by the interaction between the viral Spike protein and the human ACE2 receptor. This mechanism evolved from the ancestor bat coronavirus and is currently one of the main targets for antiviral strategies. However, there currently exist several Spike protein variants in the SARS-CoV-2 population as the result of mutations, and it is unclear if these variants may exert a specific effect on the affinity with ACE2 which, in turn, is also characterized by multiple alleles in the human population. In the current study, the GBPM analysis, originally developed for highlighting host-guest interaction features, has been applied to define the key amino acids responsible for the Spike/ACE2 molecular recognition, using four different crystallographic structures. Then, we intersected these structural results with the current mutational status, based on more than 295,000 sequenced cases, in the SARS-CoV-2 population. We identified several Spike mutations interacting with ACE2 and mutated in at least 20 distinct patients: S477N, N439K, N501Y, Y453F, E484K, K417N, S477I and G476S. Among these, mutation N501Y in particular is one of the events characterizing SARS-CoV-2 lineage B.1.1.7, which has recently risen in frequency in Europe. We also identified five ACE2 rare variants that may affect interaction with Spike and susceptibility to infection: S19P, E37K, M82I, E329G and G352V.Communicated by Ramaswamy H. Sarma.

HRE-type genes are regulated by growth-related changes in internal oxygen concentrations during the normal development of potato (Solanum tuberosum) tubers.

The occurrence of hypoxic conditions in plants not only represents a stress condition but is also associated with the normal development and growth of many organs, leading to adaptive changes in metabolism and growth to prevent internal anoxia. Internal oxygen concentrations decrease inside growing potato tubers, due to their active metabolism and increased resistance to gas diffusion as tubers grow. In the present work, we identified three hypoxia-responsive ERF (StHRE) genes whose expression is regulated by the gradual decrease in oxygen tensions that occur when potato tubers grow larger. Increasing the external oxygen concentration counteracted the modification of StHRE expression during tuber growth, supporting the idea that the actual oxygen levels inside the organs, rather than development itself, are responsible for the regulation of StHRE genes. We identified several sugar metabolism-related genes co-regulated with StHRE genes during tuber development and possibly involved in starch accumulation. All together, our data suggest a possible role for low oxygen in the regulation of sugar metabolism in the potato tuber, similar to what happens in storage tissues during seed development.

The Transcriptome of SH-SY5Y at Single-Cell Resolution: A CITE-Seq Data Analysis Workflow.

Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) is a recently established multimodal single cell analysis technique combining the immunophenotyping capabilities of antibody labeling and cell sorting with the resolution of single-cell RNA sequencing (scRNA-seq). By simply adding a 12-bp nucleotide barcode to antibodies (cell hashing), CITE-seq can be used to sequence antibody-bound tags alongside the cellular mRNA, thus reducing costs of scRNA-seq by performing it at the same time on multiple barcoded samples in a single run. Here, we illustrate an ideal CITE-seq data analysis workflow by characterizing the transcriptome of SH-SY5Y neuroblastoma cell line, a widely used model to study neuronal function and differentiation. We obtained transcriptomes from a total of 2879 single cells, measuring an average of 1600 genes/cell. Along with standard scRNA-seq data handling procedures, such as quality checks and cell filtering procedures, we performed exploratory analyses to identify most stable genes to be possibly used as reference housekeeping genes in qPCR experiments. We also illustrate how to use some popular R packages to investigate cell heterogeneity in scRNA-seq data, namely Seurat, Monocle, and slalom. Both the CITE-seq dataset and the code used to analyze it are freely shared and fully reusable for future research.

Single-Cell Gene Network Analysis and Transcriptional Landscape of MYCN-Amplified Neuroblastoma Cell Lines.

Neuroblastoma (NBL) is a pediatric cancer responsible for more than 15% of cancer deaths in children, with 800 new cases each year in the United States alone. Genomic amplification of the MYC oncogene family member MYCN characterizes a subset of high-risk pediatric neuroblastomas. Several cellular models have been implemented to study this disease over the years. Two of these, SK-N-BE-2-C (BE2C) and Kelly, are amongst the most used worldwide as models of MYCN-Amplified human NBL. Here, we provide a transcriptome-wide quantitative measurement of gene expression and transcriptional network activity in BE2C and Kelly cell lines at an unprecedented single-cell resolution. We obtained 1105 Kelly and 962 BE2C unsynchronized cells, with an average number of mapped reads/cell of roughly 38,000. The single-cell data recapitulate gene expression signatures previously generated from bulk RNA-Seq. We highlight low variance for commonly used housekeeping genes between different cells (ACTB, B2M and GAPDH), while showing higher than expected variance for metallothionein transcripts in Kelly cells. The high number of samples, despite the relatively low read coverage of single cells, allowed for robust pathway enrichment analysis and master regulator analysis (MRA), both of which highlight the more mesenchymal nature of BE2C cells as compared to Kelly cells, and the upregulation of TWIST1 and DNAJC1 transcriptional networks. We further defined master regulators at the single cell level and showed that MYCN is not constantly active or expressed within Kelly and BE2C cells, independently of cell cycle phase. The dataset, alongside a detailed and commented programming protocol to analyze it, is fully shared and reusable.