Researchers at the Crick are tackling the big questions about human health and disease, and new findings are published every week. Our faculty have picked some of the most significant papers published by Crick scientists, all of which are freely available thanks to our open science policy.
Functional cross-talk between allosteric effects of activating and inhibiting ligands underlies PKM2 regulation
This work reveals that amino acids, rather than fructose 1,6-bisphosphate, are the relevant cellular regulators of pyruvate kinase M2 (PKM2), a critical node in cancer metabolism. It further elucidates the molecular mechanism of PKM2 regulation by amino acids with a new algorithm that predicts allosteric pathways in proteins, a major and difficult problem in structural biology.
CD1d-mediated lipid presentation by CD11c+ cells regulates intestinal homeostasis
Intestinal homeostasis requires a continuous dialogue between commensal bacteria and intestinal immune cells. Natural Killer T (NKT) cells are a population of CD1d-restricted lipid-reactive lymphocytes contributing to the regulation of mucosal immunity, but the mechanisms underlying this are poorly understood. Here we show that lipid presentation by CD1d+ intestinal dendritic cells and macrophages controls NKT cell function and activation which in turn regulates commensal bacteria and immune cell populations in the gut. These results reveal an NKT cell-dendritic cell crosstalk as a key mechanism for the regulation of intestinal homeostasis.
CD9 identifies pancreatic cancer stem cells and modulates glutamine metabolism to fuel tumour growth
This work identifies a cancer stem cell (CSC) population in pancreatic ductal adenocarcinoma (PDAC) marked by the tetraspanin CD9. We showed that CD9Hi CSCs are required for the epithelial and mesenchymal cellular heterogeneity seen in PDAC. We found that CD9 assembles a protein complex involved in regulating PDAC metabolism on the cell surface. CD9 depletion dramatically inhibited PDAC growth, identifying CD9 as a therapeutic PDAC target. These findings suggest that the cellular composition of pancreatic cancer is controlled by a CSC population.
Tissue curvature and apicobasal mechanical tension imbalance instruct cancer morphogenesis
This study introduces a new technique, FLASH, which enables immunostaining of whole organs for imaging and opens up the possibility of analysing a plethora of antigens and tissues that were previously impossible to study in 3D. By achieving this feat, we were able to study epithelial deformation from the moment of transformation within the intact pancreas, to show that early tumours adopt different shapes depending on tissue curvature, due to the distribution of intracellular forces. The work connects cell mechanics with the biology of tumour development in an unprecedented manner.
A protease cascade regulates release of the human malaria parasite Plasmodium falciparum from host red blood cells
This study showed that egress involves an enzyme cascade in which the serine protease SUB1 activates a second, cysteine protease called SERA6, enabling SERA6 to rapidly and precisely cleave the major red cell cytoskeletal protein β-spectrin and dismantle the cytoskeleton. It provides the first plausible model to explain how the parasite accomplishes timely rupture of its host cell membrane.
Stabilization of reversed replication forks by telomerase drives telomere catastrophe
This study defined the mechanism leading to critically short telomeres in the absence of RTEL1 and showed that telomerase, which extends telomeres in normal cells, is pathological when forks encounter an obstacle within the telomere. We showed that replication forks stall and reverse at persistent t-loops, which creates a pseudo-telomere substrate that is inappropriately stabilised by telomerase. Removing telomerase or blocking replication fork reversal rescued telomere dysfunction in Rtel1 deficient cells. We proposed that when persistent t-loops stall the replisome, telomerase inhibits fork restart, triggering the excision of the t-loop by SLX1/4 and loss of a substantial part of the telomere.
CDK phosphorylation of TRF2 controls t-loop dynamics during the cell cycle
Evidence suggested that the telomere adopts a lasso-like t-loop configuration, which safeguards chromosome ends from being recognised as DNA double strand breaks. However, the regulation and physiological importance of t-loops in end-protection was uncertain. This study uncovered a phospho-switch in TRF2 that coordinates the timely assembly and disassembly of t-loops during the cell cycle, which protects telomeres from replication stress and an unscheduled DNA damage response. These results were the first to definitively establish the t-loop as a physiologically important structure required to suppress checkpoint activation at telomere ends.
Nervous system regionalization entails axial allocation before neural differentiation
The prevailing view of neural induction in vertebrate embryos had been that cells are initially induced with anterior (forebrain) identity and then caudalising signals convert a proportion to posterior fates (spinal cord). Using chromatin accessibility, to define how cells adopt region-specific neural fates, combined with genetic and biochemical perturbations, we found that contrary to the established model, cells commit to a regional identity before acquiring neural identity. These findings prompt a revision to textbook models of neural induction. The study illustrates our adoption of new genomic methods (ATACseq) to address long-standing questions, and our capacity to productively collaborate with computational biologists.
Patient-specific cancer genes contribute to recurrently perturbed pathways and establish therapeutic vulnerabilities in esophageal adenocarcinoma
Oesophageal adenocarcinoma shows high genetic heterogeneity making the identification of cancer drivers challenging. We developed a machine learning algorithm to identify cancer drivers in 261 oesophageal adenocarcinomas. Although most predicted drivers were rare or patient-specific, they all perturbed well-known cancer pathways. Using the recurrence of the same pathway perturbations rather than individual genes, we stratified patients into six groups different for their clinical features. We validated experimentally the contribution of these genes to disease progression and revealed acquired dependencies exploitable in therapy. This study described a new way to identify cancer drivers that we have recently further developed for application in precision oncology.
Mechanism of head-to-head MCM double-hexamer formation revealed by cryo-EM
The MCM replicative helicase is loaded onto duplex DNA as a double hexamer. Here we use time-resolved cryo-EM to show that ORC binds to its high affinity binding site to load the first MCM hexamer. ORC then releases this site and it, or another ORC molecule then binds the B2 element, which contains a degenerate ORC binding site. This binding is stabilised by a novel interaction between the Orc6 subunit of ORC and the N-terminus of the MCM hexamer. ORC then recruits and loads the second hexamer by the same mechanism as the first hexamer. We employed newly developed in silico reconstitution approaches to describe the full context of the helicase loading reaction, studied on a near-native, chromatinised origin of replication. This study radically changes our approach to investigating chromosome replication with cryo-EM.
The mechanism of eukaryotic CMG helicase activation
This paper provided the first view of how the inactive MCM double hexamer is converted to two active CMG helicases. We showed MCM remains bound to ADP after loading; firing factors trigger ADP-ATP exchange; ATP rebinding causes double hexamer splitting, initial DNA melting and CMG formation. Active helicases then translocate N-terminus first.
RAC1P29S induces a mesenchymal phenotypic switch via serum response factor to promote melanoma development and therapy resistance
Metastatic melanoma is a lethal disease, in part because of rapid acquisition of resistance to therapy. Using genetically engineered mouse models, we demonstrate that the activating RAC1 P29S mutation, present in up to 5% of melanoma patients, cooperates with BRAF as a driver of melanoma initiation and promotes BRAF inhibitor resistance. The critical RAC1 effector pathway in melanoma is shown to be the transcription factor complex SRF/MRTF, which initiates a switch to a mesenchymal-like state characterized by therapy resistance. Therapeutic targeting of SRF/MRTF may have potential to reverse BRAF inhibitor resistance in melanoma patients bearing the oncogenic RAC1 P29S mutation
Development of combination therapies to maximize the impact of KRAS-G12C inhibitors in lung cancer
KRAS is the most commonly mutated oncogene in human lung cancer, but direct targeting of RAS proteins has proved difficult. A recently developed inhibitor of G12C mutant KRAS protein inhibits lung cancer progression in mouse models but does not provide durable regressions. By studying signalling pathways required for survival of KRAS mutant cells, we demonstrate a strong and selective potentiation of the effects of G12C KRAS inhibitors when mTOR and/or IGF1R are also inhibited. Using mutant specific G12C KRAS inhibitors rather than MEK inhibitors in these combinations is associated with greater specificity and lower toxicity. We propose that adding IGF1R and mTOR inhibitors will increase the impact of G12C KRAS inhibitors in clinical trials.
α-synuclein oligomers interact with ATP synthase and open the permeability transition pore in Parkinson’s disease
Protein aggregation drives neuronal death in Parkinson’s disease, although how transition of monomeric protein structures to aggregated forms causes toxicity is unknown. We demonstrate that aggregation of the protein α-synuclein generates beta sheet-rich oligomers, which localise to the mitochondrial inner membrane, where they impair complex I-dependent respiration, induce oxidation of ATP synthase and cause mitochondrial lipid peroxidation. These oxidation events result in opening of the permeability transition pore, triggering mitochondrial swelling, and ultimately cell death. This work highlights how structural conversion of a protein changes its physiological interaction with proteins and lipids, and induces pathology in human cell models of disease.
A cell-size threshold limits cell polarity and asymmetric division potential
A key requirement for patterning networks is that the scale of pattern be appropriately matched to the size of the system to be patterned. Through a combination of theory and experiment, we show that failure of the PAR network to scale with cell size restricts stable cell polarity to a specific size range and imposes a minimum cell size threshold for polarity. Experimental alteration of cell size indicates that embryos are sensitive to this size threshold. We thus propose a general strategy by which cells can use intrinsic length scales of patterning networks to enable size-dependent decision making.
Early-life exposure to low-dose oxidants can increase longevity via microbiome remodelling in Drosophila
This reports the first identification, in any species, of the microbiome as a key mediator of developmental stress-induced longevity. We found that mild oxidative stress during development robustly increases lifespan via the selective elimination of Acetobacter from the microbiome. This study also highlights that targeted remodelling of the early-life microbiome can provide an efficient strategy for extending healthspan and lifespan.
Subcellular antibiotic visualization reveals a dynamic drug reservoir in infected macrophages
Improving chemotherapies against intracellular pathogens requires an understanding of how antibiotic distribution within infected cells affects efficacy. In this work, we developed an approach to visualise antibiotics in human macrophages infected with the tubercle bacillus. We showed that the antitubercular (anti-TB) drug bedaquiline accumulated in host lipid droplets, which seemed to act as an antibiotic reservoir that could be transferred to bacteria during host lipid consumption. Indeed, alterations in host lipid droplet content affected the anti-TB activity of bedaquiline against intracellular bacilli.
Long-range signaling activation and local inhibition separate the mesoderm and endoderm lineages
The induction of endoderm and mesoderm by the signalling molecule Nodal has long been a textbook example of how a morphogen patterns vertebrate tissues. This study overturned the view that tissues are patterned through a single long-range morphogen gradient. Instead we demonstrated that Nodal functions in an incoherent feedforward loop with Fgf, to determine endoderm and mesoderm specification. Nodal induces long-range Fgf signaling, which is required for mesoderm induction, while simultaneously inducing a cell-autonomous Fgf signaling inhibitor within cells destined to become endoderm. This work represents a major step forward in deciphering the organising principles underlying early embryonic patterning.
LTR retroelement expansion of the human cancer transcriptome and immunopeptidome revealed by de novo transcript assembly
We assembled and disseminated the most complete, to date, transcriptome with a focus on transcripts initiated by or overlapping with endogenous retroelements. This assembly doubles the number of known transcripts and forms the basis for in-depth analysis of retroelement studies in health and disease, particularly in cancer. It also provided unconventional targets for novel cancer vaccines that are being developed by Enara Bio.
Plasmodium-specific atypical memory B cells are short-lived activated B cells
This paper provides strong evidence that “atypical” B cells are short-lived activated B cells, and are probably the result of chronic stimulation and not the cause of chronic malaria. This questions the commonly held view that atypical B cells are evidence of an aberrant or defective response in malaria.