5683099 research outputs found
Sort by
Single Molecule Localisation Microscopyand Image Analysis to investigatethe sarcomere at the nanoscale
No full textThe sarcomereThe sarcomere is the base contractile unit of muscle. With a length of ~2μm, interesting sub-sarcomeric structure is at the nanoscale and super-resolution (SR) microscopy is needed to image it and understand it. Electron Microscopy is limited in the denser regions made up of very similar domains; hence SR fluorescence microscopy provides nanoscale resolution imaging with high molecular specificity labelling. SMLM is at the forefront of SR-fluorescence microscopy, and is therefore an important tool for resolving substructure at the nanoscale.Poster presented as part of the Crick BioImage Analysis Symposium 2025.Permission has been given by authors to upload to Crick Figshare. Copyright remains with the original authors.</p
Progressive chromosome shape changes during cell divisions.
No full textMitotic chromosomes give genome portions the required compaction and mechanical stability for faithful inheritance during cell divisions. They are shaped by the chromosomal condensin complex. Here, we record human chromosome dimensions from their appearance in prophase over successive times in a mitotic arrest. Chromosomes first appear long and uniformly thin. Then, individual chromosome arms become discernible, which continuously shorten and thicken-the longer a chromosome arm, the thicker it becomes. In the search for a molecular explanation of this behavior, given uniform condensin density, the popular loop extrusion model provides no obvious means by which longer chromosome arms become thicker. Instead, we find that simulations of an alternative loop capture model recapitulate key features of our observations, with re-arranging chromatin rosettes underpinning the gradually developing arm length-to-width relationship. Our analyses portray chromosomes as out-of-equilibrium structures in the process of transitioning towards, but on biologically relevant time scales not typically reaching, steady state
Marsupial single-cell transcriptomics identifies temporal diversity in mammalian developmental programs.
No full textSingle-cell transcriptomics has demonstrated conserved and divergent programs of organogenesis in mammals, but existing studies have focused on eutherians. Marsupials exhibit short gestation and complete development externally, necessitating accelerated differentiation of anterior features required for locomotion and feeding. Hence, they represent a unique outgroup for studying temporal shifts in development, known as heterochrony. Here, we generate a single-cell transcriptomic atlas of gastrulation and early organogenesis in a marsupial, the opossum Monodelphis domestica. We identify previously undocumented tissues undergoing heterochrony and find that transcriptional programs that form anterior structures initiate earlier and progress faster relative to eutherians. The result is uncoupling of transcriptional and morphological timelines, revealing unforeseen diversity in mammalian developmental sequences. Using our transcriptomic dataset, we identified translation as a candidate control mechanism by which anterior prioritization is achieved. Our findings provide insight into the asynchronous progression of developmental programs in marsupials
Investigating morphogen and patterning dynamics with optogenetic control of morphogen production.
No full textMorphogen gradients provide the patterning cues that instruct cell fate decisions during development. Here, we establish an optogenetic system for the precise spatiotemporal control in vitro of Sonic hedgehog (Shh) morphogen production. Using a tunable light-inducible gene expression system, we generate long-range Shh gradients that pattern mouse neural progenitors into spatially distinct domains, mimicking neural tube development. We investigate how biochemical features of Shh and Shh-interacting proteins affect patterning length scales. By measuring clearance rates, we determine that Shh has an extracellular half-life below 1.5 h, substantially shorter than downstream gene expression dynamics, indicating gradients are continually renewed during patterning. We provide evidence that progenitor identity acquisition and maintenance depend on both Shh concentration and exposure duration. Together, this approach provides a quantitative framework for investigating morphogen patterning, enabling reproducible control of morphogen dynamics to dissect the interplay between biochemical cues, gradient formation biophysics, and transcriptional programs underlying developmental patterning
Quantifying the zoonotic risk profile of European influenza A viruses in swine from 2010 to 2020 inclusive
No full textH1 and H3 influenza A viruses (IAVs) circulating in European pigs are markedly distinct from those circulating in other global swine populations. These viruses exhibit significant genetic diversity, further expanded by periodic interspecies transmission of IAVs from humans into pigs, followed by sustained circulation. Several zoonotic IAV infections in humans in Europe have been associated with the 1C lineage of H1 IAVs. Given the predominance of H1 detections in pigs and their zoonotic potential, we quantified antigenic evolution of H1 viruses in European pigs using ferret and pig models and assessed diversity relative to swine IAV vaccine strains. Ferret and swine antisera comparisons revealed no significant differences in antibody responses. Viruses of the 1A.3.3.2 clade exhibited reduced cross-reactivity to human seasonal vaccine strains from 2009. Viruses of the 1B.1.2.2 clade showed no cross-reactivity to the 1978 human seasonal influenza viruses nor to candidate vaccine viruses (CVVs). Clades 1C.2.1 and 1C.2.2 human variant strains had variable cross-reactivity to the tested 1C lineage CVVs, and 1C.2.4 and 1C.2.5 clade viruses exhibited rapid genetic diversification. Many viruses tested were antigenically distant from swine influenza vaccine-representative strains, highlighting the need for updated vaccine formulations. Importantly, age-stratified human serum panels revealed limited population cross-protection to tested viruses, particularly for antigenically heterogenous viruses. These findings quantify the genetic and antigenic diversity of co-circulating IAV lineages and identify specific groups of viruses that may represent a greater risk to animal and public health. These results can be used to inform future pre-pandemic preparedness efforts.
IMPORTANCE
Our data demonstrate the importance of matching swine influenza vaccine seed strains to contemporary circulating viruses and highlight how vaccine mismatch may drive additional antigenic evolution and demonstrate the need for constant monitoring and surveillance of swine viruses for the benefit of both animal and global health. These findings can be used to inform and prioritize future pre-pandemic preparedness efforts
Is health, growth and development impaired in children who are Hepatitis B-exposed but uninfected?
No full textAn estimated 254 million people are living with chronic hepatitis B virus (HBV) infection worldwide. Many infants are born to mothers with HBV but do not themselves acquire the infection. It is unclear whether this exposure to HBV in early life - without the development of active infection - may be associated with adverse outcomes. We propose the term "HBV-exposed uninfected (HBEU)", drawing parallels with the HIV field which recognises that children who are HIV-exposed but uninfected face an increased risk of adverse health outcomes. This paper explores the potential health consequences for children HBEU. We summarise existing evidence reporting on children HBEU, and also review existing knowledge from the HIV field that could inform insights. We hypothesise that children HBEU may be at increased risk of preterm birth, and/or impaired growth and neurodevelopmental delay, but comprehensive, longitudinal studies are currently lacking to support this. We propose a conceptual framework to hypothesise how exposure to HBV could potentially lead to adverse growth and neurodevelopment through both HBV-specific and universal pathways, and review the available evidence and research gaps. Data are needed to establish whether short- and long-term sequelae exist for children HBEU, and to inform evidence-based interventions to mitigate against detrimental outcomes. Establishing a comprehensive understanding of the long-term trajectory of health and well-being among children HBEU throughout childhood into adolescence will require longitudinal observational studies with appropriate control groups to characterise outcomes, identify risk factors and explore underlying mechanistic pathways
Convergent mechanisms of epithelial cell structure manipulation by intestinal pathogens.
No full textThe epithelial layer that lines the digestive system serves as the primary barrier to infection by intestinal pathogens. While this layer has evolved complex molecular mechanisms to identify and respond to infection, pathogens have also evolved equally complex mechanisms to subvert this response and remodel the epithelium to their benefit. The structure of the intestinal epithelial cell is a common target of this remodeling effort. This review focuses specifically on the phenotypes and mechanisms of epithelial cell structure manipulation that have convergently evolved in human intestinal pathogens
mTOR pathway diseases: challenges and opportunities from bench to bedside and the mTOR node.
No full textMechanistic target of rapamycin (mTOR) is a highly conserved serine/threonine kinase that regulates key cellular processes including cell growth, autophagy and metabolism. Hyperactivation of the mTOR pathway causes a group of rare and ultrarare genetic diseases. mTOR pathway diseases have diverse clinical manifestations that are managed by distinct medical disciplines but share a common underlying molecular basis. There is a now a deep understanding of the molecular underpinning that regulates the mTOR pathway but effective treatments for most mTOR pathway diseases are lacking. Translating scientific knowledge into clinical applications to benefit the unmet clinical needs of patients is a major challenge common to many rare diseases. In this article we expound how mTOR pathway diseases provide an opportunity to coordinate basic and translational disease research across the group, together with industry, medical research foundations, charities and patient groups, by pooling expertise and driving progress to benefit patients. We outline the germline and somatic mutations in the mTOR pathway that cause rare diseases and summarise the prevalence, genetic basis, clinical manifestations, pathophysiology and current treatments for each disease in this group. We describe the challenges and opportunities for progress in elucidating the underlying mechanisms, improving diagnosis and prognosis, as well as the development and approval of new therapies for mTOR pathway diseases. We illustrate the crucial role of patient public involvement and engagement in rare disease and mTOR pathway disease research. Finally, we explain how the mTOR Pathway Diseases node, part of the Research Disease Research UK Platform, will address these challenges to improve the understanding, diagnosis and treatment of mTOR pathway diseases
Autophagy machinery as exploited by viruses.
No full textViruses adapt and modulate cellular pathways to allow their replication in host cells. The catabolic pathway of macroautophagy, for simplicity referred to as autophagy, is no exception. In this review, we discuss anti-viral functions of both autophagy and select components of the autophagy machinery, and how viruses have evaded them. Some viruses use the membrane remodeling ability of the autophagy machinery to build their replication compartments in the cytosol or efficiently egress from cells in a non-lytic fashion. Some of the autophagy machinery components and their remodeled membranes can even be found in viral particles as envelopes or single membranes around virus packages that protect them during spreading and transmission. Therefore, studies on autophagy regulation by viral infections can reveal functions of the autophagy machinery beyond lysosomal degradation of cytosolic constituents. Furthermore, they can also pinpoint molecular interactions with which the autophagy machinery can most efficiently be manipulated, and this may be relevant to develop effective disease treatments based on autophagy modulation
High-throughput screening of human genetic variants by pooled prime editing.
No full textMultiplexed assays of variant effect (MAVEs) enable scalable functional assessment of human genetic variants. However, established MAVEs are limited by exogenous expression of variants or constraints of genome editing. Here, we introduce a pooled prime editing (PE) platform to scalably assay variants in their endogenous context. We first improve efficiency of PE in HAP1 cells, defining optimal prime editing guide RNA (pegRNA) designs and establishing enrichment of edited cells via co-selection. We next demonstrate negative selection screening by testing over 7,500 pegRNAs targeting SMARCB1 and observing depletion of efficiently installed loss-of-function (LoF) variants. We then screen for LoF variants in MLH1 via 6-thioguanine selection, testing 65.3% of all possible SNVs in a 200-bp region including exon 10 and 362 non-coding variants from ClinVar spanning a 60-kb region. The platform's overall accuracy for discriminating pathogenic variants indicates that it will be highly valuable for identifying new variants underlying diverse human phenotypes across large genomic regions