A5036733963- Cell death mechanisms and regulation
- Trace Elements in Health
- Autophagy in Disease and Therapy
- Mitochondrial Function and Pathology
- Phagocytosis and Immune Regulation
- Liver physiology and pathology
- Heavy Metal Exposure and Toxicity
- RNA Interference and Gene Delivery
- Ubiquitin and proteasome pathways
- ATP Synthase and ATPases Research
- RNA Research and Splicing
- RNA and protein synthesis mechanisms
- interferon and immune responses
- Heavy metals in environment
- Protein Kinase Regulation and GTPase Signaling
- Neuroscience and Neuropharmacology Research
- Occupational and environmental lung diseases
- Metabolism and Genetic Disorders
- DNA Repair Mechanisms
- Endoplasmic Reticulum Stress and Disease
- Glycosylation and Glycoproteins Research
- Ion channel regulation and function
- Environmental Toxicology and Ecotoxicology
- RNA modifications and cancer
- Advanced Proteomics Techniques and Applications
MRC Toxicology Unit
2014-2023
University of Cambridge
2018-2023
University of Leicester
2009-2021
Medical Research Council
1997-2019
Kansas City VA Medical Center
2007-2010
Trinity College Dublin
2004
Sidney Kimmel Cancer Center
1997
University of Warwick
1974-1977
The intracellular regulation of cell death pathways by cIAPs has been enigmatic. Here we show that loss promotes the spontaneous formation an platform activates either apoptosis or necroptosis. This 2 MDa complex designate "Ripoptosome" is necessary but not sufficient for death. It contains RIP1, FADD, caspase-8, caspase-10, and caspase inhibitor cFLIP isoforms. cFLIP(L) prevents Ripoptosome formation, whereas, intriguingly, cFLIP(S) assembly. When are absent, activity "rheostat" controlled...
Formation of the death-inducing signaling complex (DISC) is a critical step in death receptor-mediated apoptosis, yet mechanisms underlying assembly this key multiprotein remain unclear. Using quantitative mass spectrometry, we have delineated stoichiometry native TRAIL DISC. While current models suggest that core DISC components are present at ratio 1:1, our data indicate FADD substoichiometric relative to TRAIL-Rs or DED-only proteins; strikingly, there up 9-fold more caspase-8 than...
The death-inducing signaling complex (DISC) initiates death receptor-induced apoptosis. DISC assembly and activation are controlled by c-FLIP isoforms, which function as pro-apoptotic (c-FLIPL only) or anti-apoptotic (c-FLIPL/c-FLIPS) regulators of procaspase-8 activation. Current models assume that directly competes with for recruitment to FADD. Using a functional reconstituted DISC, structure-guided mutagenesis, quantitative LC-MS/MS, we show c-FLIPL/S binding the is instead co-operative...
Abstract Cellular senescence is a terminal differentiation state that has been proposed to have role in both tumour suppression and ageing. This view supported by the fact accumulation of senescent cells can be observed response oncogenic stress as well result normal organismal Thus, identifying vivo vitro an important diagnostic therapeutic potential. The molecular pathways involved triggering and/or maintaining phenotype are not fully understood. As consequence, markers currently utilized...
Apaf-1, by binding to and activating caspase-9, plays a critical role in apoptosis. Oligomerization of the presence dATP cytochrome <i>c</i>, is required for activation caspase-9 produces caspase apoptosome complex. Reconstitution studies with recombinant proteins have indicated that size this complex very large order ∼1.4 MDa. We now demonstrate cell lysates results formation two Apaf-1-containing complexes <i>M</i> <sub>r</sub> values MDa ∼700 kDa. Kinetic analysis demonstrates <i>in...
In mammals, apoptotic protease-activating factor 1 (Apaf-1), cytochrome c, and dATP activate caspase-9, which initiates the postmitochondrial-mediated caspase cascade by proteolytic cleavage/activation of effector caspases to form active approximately 60-kDa heterotetramers. We now demonstrate that activation either in cells or following cell lysates results formation two large but different sized protein complexes, "aposome" "microaposome". Surprisingly, most DEVDase activity lysate was...
In many forms of apoptosis, cytochrome c released from mitochondria induces the oligomerization Apaf-1 to form a caspase-activating apoptosome complex. Activation lysates in vitro with dATP and cresults formation an active caspase-processing ∼700-kDa complex, which predominates apoptotic cells, relatively inactive ∼1.4-MDa We now demonstrate that assembly complex is suppressed by normal intracellular concentrations K+. Using defined reconstitution system recombinant c, K+ also inhibits...
The long-term health risks of nanoparticles remain poorly understood, which is a serious concern given their prevalence in the environment from increased industrial and domestic use. extent to such compounds contribute cellular toxicity unclear, although it known that induction oxidative stress pathways associated with this process, proteins metabolic involved nanoparticle-mediated are largely unknown. To investigate problem further, effect TiO2 on HaCaT human keratinocyte cell line was...
Decreased autophagy contributes to malignancies, however it is unclear how impacts on tumour growth. Acute myeloid leukemia (AML) an ideal model address this as (i) patient samples are easily accessible, (ii) the hematopoietic stem and progenitor population (HSPC) where transformation occurs well characterized, (iii) loss of key gene Atg7 in cells (HSPCs) leads a lethal pre-leukemic phenotype mice. Here we demonstrate that Atg5 results identical HSPC Atg7, confirming general role for...
Mitochondrial toxicity is increasingly being implicated as a contributing factor to many xenobiotic-induced organ toxicities, including skeletal muscle toxicity. This has necessitated the need for predictive in vitro models that are able sensitively detect mitochondrial of chemical entities early research and development process. One such cell model involves substituting galactose glucose culture media. Since cells cultured unable generate sufficient ATP from glycolysis they forced rely on...
During the translation surveillance mechanism known as ribosome-associated quality control, ASC-1 complex (ASCC) disassembles ribosomes stalled on mRNA. Here, we show that there are two distinct classes of ribosome. Ribosomes by elongation inhibitors or methylated mRNA short lived in human cells because they split ASCC. In contrast, although ultraviolet light and 4-nitroquinoline 1-oxide induce ribosome stalling damaging mRNA, ASCC is recruited to these ribosomes, found refractory...
Identification of the processing/activation multiple interleukin-1beta converting enzyme (ICE)-like proteases and their target substrates in intact cell is critical to our understanding apoptotic process. In this study we demonstrate at least four ICE-like during execution phase apoptosis human monocytic tumor THP.1 cells. Apoptosis was accompanied by processing Ich-1, CPP32, Mch3alpha catalytically active subunits, lysates from these cells displayed a proteolytic activity with kinetics,...
The apoptosome is a large caspase-activating ( approximately 700-1400 kDa) complex, which assembled from Apaf-1 and caspase-9 when cytochrome c released during mitochondrial-dependent apoptotic cell death. the core scaffold protein 135 kDa contains CARD (caspase recruitment domain), CED-4, multiple (13) WD40 repeat domains, can potentially interact with variety of unknown regulatory proteins. To identify such proteins we activated THP.1 lysates dATP/cytochrome used sucrose density...
Repair of double-stranded DNA breaks (DSBs) in mammalian cells primarily occurs by the non-homologous end-joining (NHEJ) pathway, which requires seven core proteins (Ku70/Ku86, DNA-PKcs (DNA-dependent protein kinase catalytic subunit), Artemis, XRCC4-like factor (XLF), XRCC4 and ligase IV). Here we show using combined affinity purification mass spectrometry that co-purifies with all known NHEJ factors. Furthermore, have identified a novel evolutionary conserved associated DNA-PKcs-c9orf142....
Abstract Background Regulation of the mRNA life cycle is central to gene expression control and determination cell fate. miRNAs represent a critical regulatory mechanism, but despite decades research, their mode action still not fully understood. Results Here, we show that eIF4A2 major effector repressive miRNA pathway functioning via Ccr4-Not complex. We demonstrate while DDX6 interacts with Ccr4-Not, its effects in mechanism are as pronounced. Through interaction complex, represses mRNAs...
Abstract Regulated cell death is essential in development and cellular homeostasis. Multi-protein platforms, including the Death-Inducing Signaling Complex (DISC), co-ordinate fate via a core FADD:Caspase-8 complex its regulatory partners, such as inhibitor c-FLIP. Here, using electron microscopy, we visualize full-length procaspase-8 with FADD. Our structural analysis now reveals how FADD-nucleated tandem effector domain (tDED) helical filament required to orientate catalytic domains,...