T lymphocyte differentiation in the steady state is characterized by high cellular turnover whereby thymocytes do not self-renew. However, if deprived of competent progenitors, the thymus can ...temporarily maintain thymopoiesis autonomously. This bears a heavy cost, because prolongation of thymus autonomy causes leukemia. Here, we show that, at an early stage, thymus autonomy relies on double-negative 3 early (DN3e) thymocytes that acquire stem-cell-like properties. Following competent progenitor deprivation, DN3e thymocytes become long lived, are required for thymus autonomy, differentiate in vivo, and include DNA-label-retaining cells. At the single-cell level, the transcriptional programs of thymopoiesis in autonomy and the steady state are similar. However, a new cell population emerges in autonomy that expresses an aberrant Notch target gene signature and bypasses the β-selection checkpoint. In summary, DN3e thymocytes have the potential to self-renew and differentiate in vivo if cell competition is impaired, but this generates atypical cells, probably the precursors of leukemia.
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•Thymus autonomy depends on DN3e thymocytes that secure T cell differentiation•DN3e in autonomy self-renew, retain DNA label, and have enhanced proliferation•Thymus autonomy preserves the hallmarks of thymopoiesis at the single-cell level•Aberrant cells with active notch that bypassed β-selection emerge in autonomy
Thymus autonomy is the capacity to produce T cells without bone marrow contribution, which has a high risk for leukemia. Paiva et al. show that thymus autonomy depends on self-renewing DN3e thymocytes. This property generates aberrant cells that are likely the predecessors of leukemia. Hence, conditions causing thymus autonomy should be avoided.
Bulk RNA-sequencing of Plasmodium spp., the causative parasite of malaria, fails to discriminate the stage-specific gene regulation of its asexual developmental blood stages. Here, we provide a ...protocol that uses single-cell RNA sequencing of FACS-sorted Plasmodium chabaudi chabaudi AS (Pcc) infected red blood cells (iRBCs) to characterize parasite stage-specific modulation of gene expression. We describe steps for infecting mice, monitoring the disease, preparing iRBCs, and single cell sequencing iRBCs. We then detail procedures for analyzing scRNA-Seq data.
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•ScRNA sequencing protocol for Plasmodium chabaudi chabaudi blood stages.•Characterization of different Plasmodium blood stages by scRNA sequencing.•Characterization of Plasmodium blood stage transitions by scRNA trajectories.•Characterization of host effects on Plasmodium blood stages by scRNA sequencing
Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
SNF1-related Kinase 1 (SnRK1) is an evolutionarily conserved protein kinase with key functions in energy management during stress responses in plants. To address a potential role of SnRK1 under ...favorable conditions, we performed a metabolomic and transcriptomic characterization of rosettes of 20-d-old Arabidopsis (Arabidopsis thaliana) plants of SnRK1 gain- and loss-of-function mutants during the regular diel cycle. Our results show that SnRK1 manipulation alters the sucrose and trehalose 6-phosphate (Tre6P) relationship, influencing how the sucrose content is translated into Tre6P accumulation and modulating the flux of carbon to the tricarboxylic acid cycle downstream of Tre6P signaling. On the other hand, daily cycles of Tre6P accumulation were accompanied by changes in SnRK1 signaling, leading to a maximum in the expression of SnRK1-induced genes at the end of the night, when Tre6P levels are lowest, and to a minimum at the end of the day, when Tre6P levels peak. The expression of SnRK1-induced genes was strongly reduced by transient Tre6P accumulation in an inducible Tre6P synthase (otsA) line, further suggesting the involvement of Tre6P in the diel oscillations in SnRK1 signaling. Transcriptional profiling of wild-type plants and SnRK1 mutants also uncovered defects that are suggestive of an iron sufficiency response and of a matching induction of sulfur acquisition and assimilation when SnRK1 is depleted. In conclusion, under favorable growth conditions, SnRK1 plays a role in sucrose homeostasis and transcriptome remodeling in autotrophic tissues and its activity is influenced by diel fluctuations in Tre6P levels.
The Mouse Genomes Project was initiated in 2009 with the goal of using next-generation sequencing technologies to catalogue molecular variation in the common laboratory mouse strains, and a selected ...set of wild-derived inbred strains. The initial sequencing and survey of sequence variation in 17 inbred strains was completed in 2011 and included comprehensive catalogue of single nucleotide polymorphisms, short insertion/deletions, larger structural variants including their fine scale architecture and landscape of transposable element variation, and genomic sites subject to post-transcriptional alteration of RNA. From this beginning, the resource has expanded significantly to include 36 fully sequenced inbred laboratory mouse strains, a refined and updated data processing pipeline, and new variation querying and data visualisation tools which are available on the project’s website (
http://www.sanger.ac.uk/resources/mouse/genomes/
). The focus of the project is now the completion of de novo assembled chromosome sequences and strain-specific gene structures for the core strains. We discuss how the assembled chromosomes will power comparative analysis, data access tools and future directions of mouse genetics.
Hypoglycemia is a clinical hallmark of severe malaria, the often-lethal outcome of Plasmodium falciparum infection. Here, we report that malaria-associated hypoglycemia emerges from a non-canonical ...resistance mechanism, whereby the infected host reduces glycemia to starve Plasmodium. This hypometabolic response is elicited by labile heme, a byproduct of hemolysis that induces illness-induced anorexia and represses hepatic glucose production. While transient repression of hepatic glucose production prevents unfettered immune-mediated inflammation, organ damage, and anemia, when sustained over time it leads to hypoglycemia, compromising host energy expenditure and adaptive thermoregulation. The latter arrests the development of asexual stages of Plasmodium via a mechanism associated with parasite mitochondrial dysfunction. In response, Plasmodium activates a transcriptional program associated with the reduction of virulence and sexual differentiation toward the generation of transmissible gametocytes. In conclusion, malaria-associated hypoglycemia represents a trade-off of a hypometabolic-based defense strategy that balances parasite virulence versus transmission.
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•Repression of hepatic gluconeogenesis by labile heme drives malarial hypoglycemia•Hypoglycemia lowers Plasmodium virulence•Malarial hypoglycemia compromises host energy metabolism and thermoregulation•Plasmodium undergoes gametocytogenesis in response to hypoglycemia
Malaria-associated hypoglycemia develops as a trade-off of a non-canonical resistance mechanism against Plasmodium infection based on restricting parasite access to glucose. In response, Plasmodium reduces its virulence in favor of transmission, an evolutionarily conserved host-pathogen cooperative metabolic behavior.
Understanding SARS-CoV-2 evolution and host immunity is critical to control COVID-19 pandemics. At the core is an arms-race between SARS-CoV-2 antibody and angiotensin-converting enzyme 2 (ACE2) ...recognition, a function of the viral protein spike. Mutations in spike impacting antibody and/or ACE2 binding are appearing worldwide, imposing the need to monitor SARS-CoV2 evolution and dynamics in the population. Determining signatures in SARS-CoV-2 that render the virus resistant to neutralizing antibodies is critical. We engineered 25 spike-pseudotyped lentiviruses containing individual and combined mutations in the spike protein, including all defining mutations in the variants of concern, to identify the effect of single and synergic amino acid substitutions in promoting immune escape. We confirmed that E484K evades antibody neutralization elicited by infection or vaccination, a capacity augmented when complemented by K417N and N501Y mutations. In silico analysis provided an explanation for E484K immune evasion. E484 frequently engages in interactions with antibodies but not with ACE2. Importantly, we identified a novel amino acid of concern, S494, which shares a similar pattern. Using the already circulating mutation S494P, we found that it reduces antibody neutralization of convalescent and post-immunization sera, particularly when combined with E484K and with mutations able to increase binding to ACE2, such as N501Y. Our analysis of synergic mutations provides a signature for hotspots for immune evasion and for targets of therapies, vaccines and diagnostics. Author summary For a SARS-CoV-2 virion to enter a cell, the spike protein displayed at its surface must be recognized by the host ACE2 receptor. Serum neutralizing antibodies, shown to develop upon natural SARS-CoV-2 infection or vaccination, bind spike protein preventing the recognition by ACE2 and, consequently, infection. However, SARS-CoV-2 virus is constantly evolving, and can acquire mutations in spike that render this protein resistant to neutralizing antibodies and make vaccines ineffective. In this paper, we tested how single and a combination of mutations naturally occurring in spike, including in variants of concern, would synergize to affect antibody neutralizing capacity. We then integrated these findings with in silico analyses of amino acids binding to ACE2 and antibodies, and distributed them in a grid as amino acids important for binding to ACE2 or antibodies, or both. We found that changes in amino such as E484 and S494, which frequently interact with antibodies but not with ACE2, promptly evolve immune escape mutants, elicited by infection or vaccination, if the mutation severely alters the binding specificity of the antibody. Our work also suggests that the combination of these mutations with others promoting ACE2 binding, such as N501Y, increases their ability to escape neutralizing-antibody responses.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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