Three-dimensional molecular cartography of human cerebral organoids revealed by double-barcoded spatial transcriptomics.

Spatially resolved transcriptomics is revolutionizing our understanding of complex tissues, but scaling these approaches to multiple tissue sections and three-dimensional tissue reconstruction remains challenging and cost prohibitive. In this work, we present a low-cost strategy for manufacturing molecularly double-barcoded DNA arrays, enabling large-scale spatially resolved transcriptomics studies. We applied this technique to spatially resolve gene expression in several human brain organoids, including the reconstruction of a three-dimensional view from multiple consecutive sections, revealing gene expression heterogeneity throughout the tissue.

Synergistic activation of RARß and RARγ nuclear receptors restores cell specialization during stem cell differentiation by hijacking RARα-controlled programs.

How cells respond to different external cues to develop along defined cell lineages to form complex tissues is a major question in systems biology. Here, we investigated the potential of retinoic acid receptor (RAR)-selective synthetic agonists to activate the gene regulatory programs driving cell specialization during nervous tissue formation from embryonic carcinoma (P19) and mouse embryonic (E14) stem cells. Specifically, we found that the synergistic activation of the RARß and RARγ by selective ligands (BMS641 or BMS961) induces cell maturation to specialized neuronal subtypes, and to astrocytes and oligodendrocyte precursors. Using RAR isotype knockout lines exposed to RAR-specific agonists, interrogated by global transcriptome landscaping and in silico modeling of transcription regulatory signal propagation, revealed major RARα-driven gene programs essential for optimal neuronal cell specialization and hijacked by the synergistic activation of the RARß and RARγ receptors. Overall, this study provides a systems biology view of the gene programs accounting for the previously observed redundancy between RARs, paving the way toward their potential use for directing cell specialization during nervous tissue formation.

A Core Transcription Regulatory Circuitry Defining Microglia Cell Identity Inferred from the Reanalysis of Multiple Human Microglia Differentiation Protocols.

Microglia, the immune cells in the brain involved in both homeostasis and injury/infection control, play a predominant role in neurodegenerative diseases. In vivo studies on microglia are limited due to the requirement of surgical intervention, which can lead to the destruction of the tissues. Over the last few years, multiple protocols-presenting a variety of strategies-have described microglia differentiation issued from human pluripotent stem cells. Herein, we have reanalyzed the transcriptomes released on six different microglia differentiation protocols and revealed a consensus core of master transcription regulatory circuitry defining microglia identity. Furthermore, we have discussed the major divergencies among the studied protocols and have provided suggestions to further enhance microglia differentiation assays.

Protocol for using MULTILAYER to reveal molecular tissue substructures from digitized spatial transcriptomes.

Spatially resolved transcriptomics (SrT) allow researchers to explore organ/tissue architecture from the angle of the gene programs involved in their molecular complexity. Here, we describe the use of MULTILAYER to reveal molecular tissue substructures from the analysis of localized transcriptomes (defined as gexels). MULTILAYER can process low- and high-resolution SrT data but also perform comparative analyses within multiple SrT readouts. For complete details on the use and execution of this protocol, please refer to Moehlin et al., 2021.

Real-time SARS-CoV-2 diagnostic and variants tracking over multiple candidates using nanopore DNA sequencing.

Since December 2019, a novel coronavirus responsible for a severe acute respiratory syndrome (SARS-CoV-2) is accountable for a major pandemic situation. The emergence of the B.1.1.7 strain, as a highly transmissible variant has accelerated the world-wide interest in tracking SARS-CoV-2 variants' occurrence. Similarly, other extremely infectious variants, were described and further others are expected to be discovered due to the long period of time on which the pandemic situation is lasting. All described SARS-CoV-2 variants present several mutations within the gene encoding the Spike protein, involved in host receptor recognition and entry into the cell. Hence, instead of sequencing the whole viral genome for variants' tracking, herein we propose to focus on the SPIKE region to increase the number of candidate samples to screen at once; an essential aspect to accelerate diagnostics, but also variants' emergence/progression surveillance. This proof of concept study accomplishes both at once, population-scale diagnostics and variants' tracking. This strategy relies on (1) the use of the portable MinION DNA sequencer; (2) a DNA barcoding and a SPIKE gene-centered variant's tracking, increasing the number of candidates per assay; and (3) a real-time diagnostics and variant's tracking monitoring thanks to our software RETIVAD. This strategy represents an optimal solution for addressing the current needs on SARS-CoV-2 progression surveillance, notably due to its affordable implementation, allowing its implantation even in remote places over the world.