Publication highlights

Researchers at the Crick have developed a CRISPR-based screening method to rapidly assess how the loss of individual Cryptosporidium genes influence parasite survival in vivo. Using this method, they examined the parasite’s pyrimidine salvage pathway and a set of leading Cryptosporidium vaccine candidates. This targeted screening method is highly versatile and will enable researchers to more rapidly expand the knowledge base for Cryptosporidium infection biology.

Researchers have shown that the Cryptosporidium parasite exports a protein into infected intestinal cells, altering the gut environment and enabling the parasite to survive and replicate. They investigated a major protein in a family of exported proteins, called microvilli protein 1 (MVP1), finding that, within the epithelial cell, it interacts with human proteins that are responsible for maintaining structure and regulating cellular projections like microvilli. One of the proteins that MVP1 interacts with, called EBP50, is crucial for stabilising pumps on the surface of the cells that bring salts in. Disrupting these pumps results in diarrhoea, so the team believes that MVP1 might be one of the key factors that drive the symptoms caused by Cryptosporidium. They also found that MVP1 interacts with the same structural proteins as the Map protein, exported by the E. Coli bacteria, which also causes microvilli elongation.

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, infects lung macrophages and subverts immune responses. In this work, researchers at the Crick developed genetically encoded probes to visualise calcium fluxes in human macrophages. By visualising calcium, they discovered that calcium is an important signal during infection that leaks from Mtb phagosomes. This calcium flux triggers a complex membrane remodelling and the association of autophagic proteins ATG8/LC3 to these membranes. They show that this membrane remodelling is important to limit the damage that Mtb inflicts in macrophage membranes and restrict Mtb infection as part the innate immune response.

Researchers at the Crick and NPL, as part of the CRUK Grand Challenges team Rosetta, have developed a multimodal imaging pipeline that extends upon the principles of correlative light, electron, and ion microscopy (CLEIM), which combines confocal microscopy reporter or probe-based fluorescence, electron microscopy (EM), stable isotope labelling and Nanoscale secondary ion mass spectrometry (NanoSIMS). Their protocol allows an unprecedented extraction of biological information from specimens, whilst being based on a series of well-established and widely available technologies, thus allowing quick adaptation of the protocol for individual research needs. This integration provides a multifaceted view of the tissue microenvironment, capturing both the internal cellular architecture and the intricate metabolic dynamics occurring within. The researchers tested their pipeline by imaging the incorporation of carbon from glucose into B and T cells in mouse liver tumours.

Scientists at the Crick have generated human stem cell models which, for the first time, contain notochord – a tissue in the developing embryo that acts like a navigation system, directing cells where to build the spine and nervous system (the trunk). The team first analysed chicken embryos to understand exactly how the notochord forms naturally. By comparing this with existing published information from mouse and monkey embryos, they established the timing and sequence of the molecular signals needed to create notochord tissue. With this blueprint, they produced a precise sequence of chemical signals and used this to coax human stem cells into forming a notochord. The stem cells formed a miniature ‘trunk-like’ structure, which spontaneously elongated to 1-2 millimetres in length. The scientists believe this work could help to study birth defects affecting the spine and spinal cord.

The Stem Cell Biology and Developmental Genetics Laboratory at the Crick has shown that in a mouse model of human hypopituitarism, low doses of aspirin and the balance of bacteria in the gut can influence symptoms. The brains of these mice, which lack the Sox3 gene, had a reduced number of a hypothalamic cell type, NG2 glial cells. Treating Sox3-deficient animals with a low dose of aspirin for 21 days increased NG2 glia numbers and reversed the hypopituitarism, but highly surprisingly, the makeup of the gut microbiome also had a significant effect; the change in microbiome resulting from the mice migrating into the Crick animal facility from elsewhere was enough to rescue the hormonal deficiencies. In addition to providing a clear case where extrinsic factors can influence a robust phenotype caused by a mutation, the work has implications for experimental consistency between research facilities, and has garnered wide interest in the field.

The human RAD52 protein plays an important role in several cellular processes, including the repair of chromosome breaks and the maintenance of telomere length (structures at the end of chromosomes) to avoid cellular aging. During DNA repair, it provides an alternative to the BRCA2 protein, which is mutated in many inheritable breast, ovarian and prostate cancers. Consequently, targeting RAD52 could be used to kill tumours with BRCA2 mutations, where growth is uncontrolled. To elucidate the mechanism of repair by RAD52, we determined the atomic structure of the protein using cryo-electron microscopy, and found that the protein forms a ring in which the broken DNA wraps around the outside of the ring. Having the atomic structure gives us new insights into ways to identify small molecules that can be used to inhibit repair by RAD52 and kill BRCA2-defective tumours.

Researchers at the Crick have now developed a way to grow specialised left ventricular heart muscle cells from stem cells, opening up new opportunities for research into heart disease, drug screening, and potentially the development of new treatments.

Their methods are published today in Cell Reports Methods and have also been licensed to Axol Bioscience to commercialise the protocol for the generation and sale of cardiomyocytes for R&D and the provision of contract research services, especially in field of drug screening and cardiotoxicity assays.