Publication highlights

A Crick researcher reading a scientific paper on a screen.

Intro

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.

Highlights

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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.

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Published in eLife

Published

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.

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Published in Nature Cell Biology

Published

3D image of a pancreas

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.

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Published in Nature

Published

Image of mouse eyeball taken with light-sheet fluorescent microscopy, with the blood vessels shown in green.

Mouse retinal cell behaviour in space and time using light sheet fluorescence microscopy

We successfully performed the first lightsheet 3D/4D imaging of mouse retinas (focussing on vessels and neurons) to demonstrate that current confocal methods distort vessel tissue. This brings a much improved way to observe and quantify the devastating changes to vessels and neurons in retinopathy mouse models.

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Published in eLife

Published

Reconstitution of a functional human thymus by postnatal stromal progenitor cells and natural whole-organ scaffolds

In this paper we define the heterogeneity and the clonogenic potential of human thymus stroma; characterise progenitor cells capable of extensive expansion in vitro, thereby achieving clinically relevant numbers with resilience to long-term storage; and report an epithelial-mesenchymal hybrid phenotype of thymus epithelial cells in vivo and in vitro that affects cell behaviour, a unique feature among any epithelia so far reported. We describe a protocol for organs that lack a main vascular access that allowed us to specify the role of natural ECM in supporting organ morphogenesis ex vivo and in vivo; and reconstitute a functional human thymus long-term in vivo.

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Published in Nature Communications

Published

Chromosomes showing telomeres at their ends.

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.

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Published in Nature

Published

Microscope images of mouse and human spinal cord development at equivalent stages.

Species-specific pace of development is associated with differences in protein stability

Despite evolutionarily conservation of molecular mechanisms, the speed of development varies substantially between species. Using in vitro directed differentiation of embryonic stem cells to motor neurons, we show that the programme of motor neuron differentiation runs twice as fast in mouse as in human. We provide evidence that a two-fold increase in protein stability and cell cycle duration in human cells compared to mouse can account for the slower pace of human development, indicating that global differences in kinetic parameters play a major role in interspecies differences in developmental tempo. This study establishes a new experimental system in which to address fundamental questions.

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Published in Science

Published

Restriction of memory B cell differentiation at the germinal center B cell positive selection stage

Memory B cells (MBCs) are key for protection from reinfection. However, it is mechanistically unclear how germinal center (GC) B cells differentiate into MBCs. MYC is transiently induced in cells fated for GC expansion and plasma cell (PC) formation, so-called positively selected GC B cells. We found that these cells coexpressed MYC and MIZ1 (MYC-interacting zinc-finger protein 1 [ZBTB17]). MYC and MIZ1 are transcriptional activators; however, they form a transcriptional repressor complex that represses MIZ1 target genes. Mice lacking MYC-MIZ1 complexes displayed impaired cell cycle entry of positively selected GC B cells and reduced GC B cell expansion and PC formation. Notably, absence of MYC-MIZ1 complexes in positively selected GC B cells led to a gene expression profile alike that of MBCs and increased MBC differentiation. Thus, at the GC positive selection stage, MYC-MIZ1 complexes are required for effective GC expansion and PC formation and to restrict MBC differentiation. We propose that MYC and MIZ1 form a module that regulates GC B cell fate.

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Published in Journal of Experimental Medicine

Published

D-Cycloserine destruction by alanine racemase and the limit of irreversible inhibition

D-cycloserine is an antibiotic used for decades to treat drug resistant tuberculosis. Its inhibition mechanism came into question when in a previous paper we determined alanine racemase activity in “fully inhibited” cells. This study demonstrated a previously unknown path during the assumed irreversible inhibition of alanine racemase that leads to the destruction of the antibiotic, meaning that alanine racemase is not irreversibly inhibited by the drug. The paper highlights the complexity of studying the chemical mechanisms of inhibition of enzymes and points to a novel strategy to design D-cycloserine analogues with improved properties.

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Published in Nature Chemical Biology

Published

Image depicting the structure of the active site of the integrase enzyme bound with the drug bictegravir.

Structural basis of second-generation HIV integrase inhibitor action and viral resistance

HIV integrase inhibitors represent some of the most impactful antimicrobial inhibitors. The second-generation drugs display improved barriers to the emergence of resistance, which spearheaded their worldwide rollout. Yet not even the most advanced compounds are immune to viral resistance. Our results explained the mechanism of viral resistance associated with the most common drug resistance mutations. Furthermore, we established the key difference between the first and second-generation strand transfer inhibitors, which will inform further development of this drug class.

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Published in Science

Published

Stochastic neighbour embedding plot.

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.

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Published in Nature Communications

Published

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.

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Published in Nature

Published

Scalable and robust SARS-CoV-2 testing in an academic center

This paper decribes how we were able to successfully repurpose the Crick to increase the capacity for Sars-CoV-2 testing in unpredented times.

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Published in Nature Biotechnology

Published

COVID testing

Pandemic peak SARS-CoV-2 infection and seroconversion rates in London frontline health-care workers

This important paper showed very high levels of infection amongst healthcare workers in a local hospital. It has influenced government policy – asymptomatic healthcare workers are to be screened as per our recommendation (announced October 12th).

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Published in The Lancet

Published

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

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Published in Cancer Cell

Published

Image showing a mouse lung before and after treatment, with tumours indicated by different colours. The tumours on the right are visibly smaller.

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.

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Published in Science Translational Medicine

Published

Modular microfluidics enables kinetic insight from time-resolved cryo-EM

Cryo-EM has the potential to study any native conformation of a macromolecule. However, the sample preparation time is high, compared to the timescale of most protein interactions and conformational changes. In this paper, we established a robust method of time-resolved cryo-EM sample preparation. We produced high-quality samples for microscopy while speeding up the process of making them by several orders of magnitude. This allowed samples to be collected within 30ms of the initiation of a biochemical reaction, within the timeframe of many critically important and interesting processes. This enables a whole new class of experiments in structural biology research.

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Published in Nature Communications

Published

Spike protein of SARS-CoV-2.

SARS-CoV-2 and bat RaTG13 spike glycoprotein structures inform on virus evolution and furin-cleavage effects

We have been able to apply the knowledge we have gained from our work on the infectivity of the influenza virus to the challenge presented by the recent SARS-CoV-2 virus outbreak. In this paper we present high resolution cryo EM structures of the SARS-CoV-2 and bat RaTG13 spike glycoproteins. We describe from a structural perspective the significant differences between the strains. We draw particular attention to the addition of a furin cleavage site into the human virus spike protein. We discuss its potential role in infectivity and on the evolution of this virulent strain.

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Published in Nature Structural & Molecular Biology

Published

Receptor binding and priming of the spike protein of SARS-CoV-2 for membrane fusion

Here we describe the conformational changes that the SARS-Cov2 spike protein undergoes in binding to the human ACE2 receptor. This represents the initial stages of the mechanism of cell invasion by the virus particle during infection. We show a series of ten cryoEM reconstructions of the spike protein binding to ACE2 through its receptor binding domain (RBD), ranging from a closed unbound spike ectodomain trimer to the fully open conformation with each RBD in the trimer bound to an ACE2 receptor. Binding to ACE2 releases the so-called fusion peptide segment and promotes membrane fusion leading to cell invasion.

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Published in Nature

Published

Alpha synuclein aggregation drives ferroptosis: an interplay of iron, calcium and lipid peroxidation

Aberrant protein-lipid interactions occur in neurodegeneration, although their role is unclear. We show how the protein α-synuclein interacts with lipids to drive a form of cell death, ferroptosis. As α-synuclein aggregates, oligomeric species with hydrophobic domains incorporate into the plasmalemmal membrane, leading to altered membrane conductance and abnormal calcium influx following glutamatergic and dopaminergic stimuli. Aggregates induce iron dependent generation of free radicals, and peroxidation of polyunsaturated fatty acids, which underlies the incorporation of aggregates into the membranes. Targeted inhibition of lipid peroxidation prevents the aggregate-membrane interaction, abolishes aberrant calcium fluxes, and restores physiological calcium signaling in human neurons, highlighting a new causative role for lipid homeostasis in Parkinson’s disease.

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Published in Cell Death and Differentiation

Published