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Scott C.C.,University of Geneva | Vossio S.,University of Geneva | Vacca F.,University of Geneva | Snijder B.,University of Zurich | And 10 more authors.
EMBO Reports | Year: 2015

The Wnt pathway, which controls crucial steps of the development and differentiation programs, has been proposed to influence lipid storage and homeostasis. In this paper, using an unbiased strategy based on high-content genome-wide RNAi screens that monitored lipid distribution and amounts, we find that Wnt3a regulates cellular cholesterol. We show that Wnt3a stimulates the production of lipid droplets and that this stimulation strictly depends on endocytosed, LDL-derived cholesterol and on functional early and late endosomes. We also show that Wnt signaling itself controls cholesterol endocytosis and flux along the endosomal pathway, which in turn modulates cellular lipid homeostasis. These results underscore the importance of endosome functions for LD formation and reveal a previously unknown regulatory mechanism of the cellular programs controlling lipid storage and endosome transport under the control of Wnt signaling. Synopsis This study reports that Wnt signaling regulates the production of lipid droplets in a process that strictly depends on endocytosed, LDL-derived cholesterol and functional endosomes. High-content screening reveals that Wnt controls cellular cholesterol homeostasis. Wnt stimulation induces accumulation of lipid droplets. Wnt mobilizes LDL-derived cholesterol through endosomes. This study reports that Wnt signaling regulates the production of lipid droplets in a process that strictly depends on endocytosed, LDL-derived cholesterol and functional endosomes. © 2015 The Authors.

O'Sullivan R.J.,Salk Institute for Biological Studies | Kubicek S.,Cambridge Broad Institute | Kubicek S.,Research Center for Molecular Medicine | Schreiber S.L.,Cambridge Broad Institute | And 3 more authors.
Nature Structural and Molecular Biology | Year: 2010

During replicative aging of primary cells morphological transformations occur, the expression pattern is altered and chromatin changes globally. Here we show that chronic damage signals, probably caused by telomere processing, affect expression of histones and lead to their depletion. We investigated the abundance and cell cycle expression of histones and histone chaperones and found defects in histone biosynthesis during replicative aging. Simultaneously, epigenetic marks were redistributed across the phases of the cell cycle and the DNA damage response (DDR) machinery was activated. The age-dependent reprogramming affected telomeric chromatin itself, which was progressively destabilized, leading to a boost of the telomere-associated DDR with each successive cell cycle. We propose a mechanism in which changes in the structural and epigenetic integrity of telomeres affect core histones and their chaperones, enforcing a self-perpetuating pathway of global epigenetic changes that ultimately leads to senescence. © 2010 Nature America, Inc. All rights reserved.

Dancik V.,The Broad Institute of MIT and Harvard | Dancik V.,Slovak Academy of Sciences | Carrel H.,The Broad Institute of MIT and Harvard | Bodycombe N.E.,The Broad Institute of MIT and Harvard | And 7 more authors.
Journal of Biomolecular Screening | Year: 2014

High-throughput screening allows rapid identification of new candidate compounds for biological probe or drug development. Here, we describe a principled method to Generate "assay performance profiles" for individual compounds that can serve as a basis for similarity searches and cluster analyses. Our method overcomes three challenges associated with Generating robust assay performance profiles: (1) we transform data, allowing us to build profiles from assays having diverse dynamic ranges and variability; (2) we apply appropriate mathematical principles to handle missing data; and (3) we mitigate the fact that loss-of-signal assay measurements may not distinguish between multiple mechanisms that can lead to certain phenotypes (e.g., cell death). Our method connected compounds with similar mechanisms of action, enabling prediction of new targets and mechanisms both for known bioactives and for compounds emerging from new screens. Furthermore, we used Bayesian modeling of promiscuous compounds to distinguish between broadly bioactive and narrowly bioactive compound communities. Several examples illustrate the utility of our method to support mechanism-of-action studies in probe development and target identification projects. ©© 2014 Society for Laboratory Automation and Screening.

Sohrabi M.,Research Center for Molecular Medicine | Mohammadi Roushandeh A.,Endometrium and Endometriosis Research Center | Alizadeh Z.,Endometrium and Endometriosis Research Center | Hosseini M.,Hamadan University of Medical Sciences
Singapore Medical Journal | Year: 2015

INTRODUCTION The aim of this study was to determine the effect of a high-fat diet (HFD) on oocyte maturation and quality in a mouse model. METHODS Female BALB/c mice were allocated to one of the following groups: (a) control group (n = 40), which received a controlled diet; or (b) HFD group (n = 40), which received an HFD for 12 weeks. Sections of the ovary were examined histologically. The number of follicles and corpora lutea were counted. In vitro maturation and in vitro fertilisation (IVF) were assessed in germinal vesicle (GV) and metaphase II (MII) oocytes, respectively. The expression of bone morphogenetic protein 15 (BMP15) and leptin receptor genes in GV and MII oocytes was evaluated using reverse transcription real-time polymerase chain reactions. RESULTS In the HFD group, there was a decreased number of primordial and Graafian follicles, as well as corpora lutea (p < 0.05). The rate of oocyte development to the MII stage was also reduced (p < 0.001). Cumulus expansion was observed more frequently in the control group than the HFD group (p < 0.05). The IVF rate in the HFD group was lower than that in the control group (p < 0.05). In the HFD group, BMP15 and leptin receptor genes were upregulated in the GV stage (p > 0.05) and MII stage (p < 0.05), compared to the control group. CONCLUSION An HFD reduces folliculogenesis in the primordial and Graafian stages, in vitro maturation and in vitro fertilisation rates, as well as oocyte quality in mice. © 2015, Singapore Medical Association. All rights reserved.

News Article
Site: cen.acs.org

The enzyme MTH1 is a hot cancer drug target. Spurred in part by a pair of 2014 Nature papers that reported MTH1 inhibitors can kill cancer cells (C&EN, April 7, 2014, page 9), more than 20 industry and academic groups have started programs to develop small molecules to shut down the enzyme. In those 2014 papers, the research teams—one led by Thomas Helleday of the Karolinska Institute and the other by Giulio Superti-Furga of CeMM, the Research Center for Molecular Medicine of the Austrian Academy of Sciences—showed that MTH1 inhibitors attack cancer cells by blocking a process that protects the cells from the effects of oxidative stress (Nature 2014, DOI: 10.1038/nature13181and 10.1038/nature13194). Cancer cells operate under oxidative stress, which can kill the cells by damaging their DNA bases. MTH1 prevents the incorporation of bases damaged by oxidation into the DNA replication process, permitting cancer cells to proliferate and thrive. However, new research suggests that blocking MTH1 may not be a deathblow to cancer cells and that the cancer-killing abilities of reported MTH1 inhibitors may not be due to shutting down the enzyme. At an American Association for Cancer Research conference last November and in a follow-up paper (Bioorg. Med. Chem. Lett. 2016, DOI: 10.1016/j.bmcl.2016.02.026), Alessia Petrocchi and coworkers at the University of Texas MD Anderson Cancer Center report a compound that potently inhibits MTH1 activity in cancer cells but doesn’t actually kill the cells. Jason G. Kettle and coworkers at Astra­Zeneca also identified small molecules that block MTH1’s enzymatic action but have limited or zero anticancer activity. When they do kill cancer cells, it’s through “off-target, nonspecific, and MTH1-independent effects on cell growth,” the researchers report (J. Med. Chem. 2016, DOI: 10.1021/acs.jmedchem.5b01760). Furthermore, the team showed that cancer cells remained viable after silencing the MTH1 gene using siRNA or CRISPR. MTH1 inhibitors still killed these cells, further supporting the conclusion that other mechanisms besides MTH1 inhibition are at play. Kettle notes that an additional unpublished study by a “leading U.K. organization also fails to validate the MTH1 mechanism.” The AstraZeneca group concludes that the enzyme’s role in cancer cells and the usefulness of inhibiting it remain uncertain. On the basis of the new findings, more careful study is now needed on MTH1’s role as a cancer drug target, says MTH1 specialist Yusaku Nakabeppu of Kyushu University. Chuan He, an expert on nucleic acid modifications at the University of Chicago, calls the siRNA knockdown and CRISPR knockout data “quite convincing” and agrees that “more work is required to assess the role of MTH1 in cancer cell survival.” But not all researchers in the field are ready to abandon MTH1 just yet. Oxidative stress expert Priyamvada Rai of the University of Miami Miller School of Medicine says, “Our independent research consistently supports MTH1 as a valid target in cancer cells that sustain oxidative stress.” Also, the anticancer abilities of molecules cannot always be investigated effectively with cell studies such as those used in the two new papers, says Jessica Martinsson, head of medicinal chemistry at Sweden’s Sprint Bioscience, which has worked on MTH1 inhibitors. Helleday and Superti-Furga still think MTH1 inhibition plays a key role in their compounds’ abilities to attack cancer. Superti-Furga wonders whether the discrepancies between the 2014 papers and the recent reports are due to intricacies in the regulation of the MTH1 pathway, which isn’t well understood. He adds that if MTH1 inhibitors do have effects in the absence of MTH1, then it will be important to find out what the additional targets are. Helleday points out that the new findings are reminiscent of recent developments on another cancer drug target, poly(ADP-ribose) polymerase (PARP). There isn’t a detailed understanding of how inhibiting the enzyme kills cancer cells, he says, but this hasn’t stopped a PARP inhibitor—AstraZeneca’s Lynparza (olaparib)—from being approved by the Food & Drug Administration for treatment of ovarian cancer and getting expedited review status for treatment of prostate cancer. “The underlying MTH1 biology is much more complex than PARP biology,” Helleday says, “so we cannot exclude as-yet-unknown interactions that could act synergistically to produce the impressive cell-killing effects we observe with our compounds. I am convinced that connecting scientists across the world using open innovation, we can solve this complex puzzle together, in time.”

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