Acetyl-CoA synthetase 2 promotes acetate utilization and maintains cancer cell growth under metabolic stress

A functional genomics study revealed that the activity of acetyl-CoA synthetase 2 (ACSS2) contributes to cancer cell growth under low-oxygen and lipid-depleted conditions. Comparative metabolomics and lipidomics demonstrated that acetate is used as a nutritional source by cancer cells in an ACSS2-dependent manner, and supplied a significant fraction of the carbon within the fatty acid and phospholipid pools. ACSS2 expression is upregulated under metabolically stressed conditions and ACSS2 silencing reduced the growth of tumor xenografts. ACSS2 exhibits copy-number gain in human breast tumors, and ACSS2 expression correlates with disease progression. These results signify a critical role for acetate consumption in the production of lipid biomass within the harsh tumor microenvironment

Melanoma cells break down LPA to establish local gradients that drive chemotactic dispersal.

The high mortality of melanoma is caused by rapid spread of cancer cells, which occurs unusually early in tumour evolution. Unlike most solid tumours, thickness rather than cytological markers or differentiation is the best guide to metastatic potential. Multiple stimuli that drive melanoma cell migration have been described, but it is not clear which are responsible for invasion, nor if chemotactic gradients exist in real tumours. In a chamber-based assay for melanoma dispersal, we find that cells migrate efficiently away from one another, even in initially homogeneous medium. This dispersal is driven by positive chemotaxis rather than chemorepulsion or contact inhibition. The principal chemoattractant, unexpectedly active across all tumour stages, is the lipid agonist lysophosphatidic acid (LPA) acting through the LPA receptor LPAR1. LPA induces chemotaxis of remarkable accuracy, and is both necessary and sufficient for chemotaxis and invasion in 2-D and 3-D assays. Growth factors, often described as tumour attractants, cause negligible chemotaxis themselves, but potentiate chemotaxis to LPA. Cells rapidly break down LPA present at substantial levels in culture medium and normal skin to generate outward-facing gradients. We measure LPA gradients across the margins of melanomas in vivo, confirming the physiological importance of our results. We conclude that LPA chemotaxis provides a strong drive for melanoma cells to invade outwards. Cells create their own gradients by acting as a sink, breaking down locally present LPA, and thus forming a gradient that is low in the tumour and high in the surrounding areas. The key step is not acquisition of sensitivity to the chemoattractant, but rather the tumour growing to break down enough LPA to form a gradient. Thus the stimulus that drives cell dispersal is not the presence of LPA itself, but the self-generated, outward-directed gradient

EXD2 Protects Stressed Replication Forks and Is Required for Cell Viability in the Absence of BRCA1/2.

Accurate DNA replication is essential to preserve genomic integrity and prevent chromosomal instability-associated diseases including cancer. Key to this process is the cells' ability to stabilize and restart stalled replication forks. Here, we show that the EXD2 nuclease is essential to this process. EXD2 recruitment to stressed forks suppresses their degradation by restraining excessive fork regression. Accordingly, EXD2 deficiency leads to fork collapse, hypersensitivity to replication inhibitors, and genomic instability. Impeding fork regression by inactivation of SMARCAL1 or removal of RECQ1's inhibition in EXD2-/- cells restores efficient fork restart and genome stability. Moreover, purified EXD2 efficiently processes substrates mimicking regressed forks. Thus, this work identifies a mechanism underpinned by EXD2's nuclease activity, by which cells balance fork regression with fork restoration to maintain genome stability. Interestingly, from a clinical perspective, we discover that EXD2's depletion is synthetic lethal with mutations in BRCA1/2, implying a non-redundant role in replication fork protection.Work in W.N.’s laboratory is funded by ICR Intramural Grant and Cancer Research UK Programme (A24881). R.A.S. and L.S. were supported by WIMM Senior Non-Clinical Fellowship awarded to W.N. M.M.S. and M.A.B. were supported by the Intramural Research Program of the NIH, National Institute on Aging, United States (Z01-AG000746-08). Work in S.G.’s laboratory is supported by BRFAA Intramural Funds. V.C. was supported by John S. Latsis Public Benefit Foundation and Alexander S. Onassis Public Benefit Foundation. Work in the P.P.’s laboratory is supported by grants from the Agence Nationale pour la Recherche (ANR), the Ligue Contre le Cancer (équipe labellisée), SIRIC Montpellier Cancer (INCa Inserm DGOS 12553), and the MSDAvenir fund

A dynamic actin cytoskeleton is required to prevent constitutive VDAC-dependent MAPK-signalling and aberrant lipid homeostasis.

The dynamic nature of the actin cytoskeleton is required to coordinate many cellular processes and a loss of its plasticity has been linked to accelerated cell ageing and attenuation of adaptive response mechanisms. Cofilin is an actin-binding protein that controls actin dynamics and has been linked to mitochondrial signalling pathways that control drug resistance and cell death. Here we show that cofilin-driven chronic depolarisation of the actin cytoskeleton activates cell wall integrity MAPK-signalling and disrupts lipid homeostasis in a VDAC-dependent manner. Expression of the cof1-5 mutation, which reduces the dynamic nature of actin, triggers loss of cell wall integrity, vacuole fragmentation, disruption of lipid homeostasis, lipid droplet (LD) accumulation and the promotion of cell death. The integrity of the actin cytoskeleton is therefore essential to maintain the fidelity of MAPK signalling, lipid homeostasis and cell health in S. cerevisiae. Graphical abstrac

Galaxy Zoo: quantitative visual morphological classifications for 48 000 galaxies from CANDELS

We present quantified visual morphologies of approximately 48 000 galaxies observed in three Hubble Space Telescope legacy fields by the Cosmic Assembly Near-infrared Deep Extragalactic Legacy Survey (CANDELS) and classified by participants in the Galaxy Zoo project. 90 per cent of galaxies have z ≤ 3 and are observed in rest-frame optical wavelengths by CANDELS. Each galaxy received an average of 40 independent classifications, which we combine into detailed morphological information on galaxy features such as clumpiness, bar instabilities, spiral structure, and merger and tidal signatures. We apply a consensus-based classifier weighting method that preserves classifier independence while effectively down-weighting significantly outlying classifications. After analysing the effect of varying image depth on reported classifications, we also provide depth-corrected classifications which both preserve the information in the deepest observations and also enable the use of classifications at comparable depths across the full survey. Comparing the Galaxy Zoo classifications to previous classifications of the same galaxies shows very good agreement; for some applications, the high number of independent classifications provided by Galaxy Zoo provides an advantage in selecting galaxies with a particular morphological profile, while in others the combination of Galaxy Zoo with other classifications is a more promising approach than using any one method alone. We combine the Galaxy Zoo classifications of ‘smooth’ galaxies with parametric morphologies to select a sample of featureless discs at 1 ≤ z ≤ 3, which may represent a dynamically warmer progenitor population to the settled disc galaxies seen at later epochs

Understanding the role of the TOPBP1 and BLM interaction in promoting genome stability

The ability to sense, respond to and repair DNA damage is essential in normal development and survival of an organism. A number of human congenital syndromes are associated with mutations in pathways involved in the DNA damage response, including Seckel and Bloom syndromes. Such diseases are often characterised by developmental abnormalities and cancer, emphasising the importance of gaining a deeper understanding of the mechanisms underlying these pathways. The protein kinase ATR, mutated in Seckel syndrome, is a critical mediator of the intra S-phase checkpoint in response to replicative stress. ATR activation is a multistep process that requires its interaction with TOPBP1. Recently published data identified an interaction between TOPBP1 and the BLM helicase. This interaction is dependent on the phosphorylation of the conserved serine 304 of BLM and mutation of this residue results in genome instability. However, exactly how TOPBP1-BLM interaction protects the genome remains unclear. The findings presented in this thesis demonstrate that DT40 cells expressing the BLM S251A mutant are defective in activation of the ATR kinase upon DNA damage. In line with this, these cells display reduced CHK1 phosphorylation, increased origin firing and unreliable replication fork restart. Importantly, these phenotypes could be rescued by fusing the ATR activating domain of TOPBP1 to the mutated BLM. Furthermore, results from BLM-deficient human cell lines demonstrate similar phenotypes. Thus, this data establishes a non-enzymatic role for the BLM helicase in promoting genome stability via activation of the ATR kinase. This may help in explaining why many Bloom syndrome patients display many of the same symptoms as ATR-Seckel patients, including short stature and microcephaly.</p

PLD2KO mice suffer from late-onset anosmia.

<p>Independent tests indicate that PLD2 KO mice have olfaction defect and that these defects are age-dependent and begin after 13 weeks of age. <b>A</b>: Buried food test; mice were timed for the retrieval of a small piece of chocolate hidden under bedding. In 72% of trials WT mice find the bait in less than 600 seconds, in contrast in only 34.8% of trials are PLD2KO mice able to find it; p<0.005. <b>B and B’</b>: Immunostaining of olfactory bulbs cryosections against c-fos (green) and counterstained with Hoechst (blue) identifies activated mitral cells in the mitral cell layer (arrow). Scale bar = 50μm. There is no difference in the number of active, c-fos positive, mitral cells between WT and PLD2KO male mice (blue and red, respectively) that have been challenged with vanilla or chocolate 30 minutes prior to sampling. <b>C</b>: Habituation-deshabituation; mice were used for three days to a cotton bud smelling of orange and on the fourth day were presented with a cotton bud smelling of vanilla. While PLD2KO mice (blue line) do not differentiate the smell, WT mice (orange line) show a regain interest in the new odour; p<0.001. <b>D</b>: The difference in habituation deshabituation response to olfactory cues is age dependent as there is no difference between WT and PLD2KO in mice that are 11 weeks of age, unlike the 14 week-old cohort presented in (C). <b>E</b>: When different objects that have the same smell are presented to the mice, WT and PLD2KO react similarly. <b>F</b>: PLD2KO mice are as good as WT mice at learning and sustaining the accelerating rotarod test.</p

The cerebellar architecture is affected in PLD2KO.

<p>A: Normally, the Purkinje cells (Pjc) form a monolayer sandwiched between the granular layer (Gr) and the molecular layer (Mol). In PLD2KO, ectopic Purkinje cells are also found either in the arbor vitae (arb, arrowhead) or clustered on the surface of the molecular layer (arrow). Scale bar = 100μm. B: PLD2KO mice have significantly more ectopic Purkinje cells than WT mice (p<0.005)</p

Lipidomic analysis indicates lipid imbalance in PLD2KO mouse brain.

<p><b>A-F</b> Lipidomic analysis of six regions of the brain indicate that PLD2KO mice have significant changes in their pools of PA. Percentage of PA species are calculated compared to the total sum of PA species identified. A) 32: species, B) 34: species, C) 36: species, D) 38: species, E) Polyunsaturated species, F) Saturated- and monounsaturated species. <i>CRB</i>, Cerebellum; <i>CTX</i>, Cortex; <i>OLF</i>, Olfactory bulbs; <i>TH</i>, Thalamus-Hypothalamus and Striatum; <i>HP</i>, Hippocampus; <i>Rest</i>, Midbrain-Hindbrain and Medulla. <b>G</b> Comparison of difference between percentage of species in WT and PLD2KO indicate that the overall proportion of 32:, 34: and 36: species have significantly changed. A-F significance is calculated by Univariate Analysis of Variance test over two different experiments. G significance is calculated as paired t-test comparing the profile of all brain regions. When p<0.05, *; p<0.01, **; p<0.005, ***</p