ATRX and RECQ5 Define Distinct Homologous Recombination Subpathways
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Prospects & Overviews Meiotic Versus Mitotic Recombination: Two Different
Prospects & Overviews Meiotic versus mitotic recombination: Two different routes for double-strand Review essays break repair The different functions of meiotic versus mitotic DSB repair are reflected in different pathway usage and different outcomes Sabrina L. Andersen1) and Jeff Sekelsky1)2)Ã Studies in the yeast Saccharomyces cerevisiae have vali- Introduction dated the major features of the double-strand break repair (DSBR) model as an accurate representation of The existence of DNA recombination was revealed by the behavior of segregating traits long before DNA was identified the pathway through which meiotic crossovers (COs) are as the bearer of genetic information. At the start of the 20th produced. This success has led to this model being century, pioneering Drosophila geneticists studied the behav- invoked to explain double-strand break (DSB) repair in ior of chromosomal ‘‘factors’’ that determined traits such as other contexts. However, most non-crossover (NCO) eye color, wing shape, and bristle length. In 1910 Thomas Hunt recombinants generated during S. cerevisiae meiosis do Morgan published the observation that the linkage relation- not arise via a DSBR pathway. Furthermore, it is becom- ships of these factors were shuffled during meiosis [1]. Building on this discovery, in 1913 A. H. Sturtevant used ing increasingly clear that DSBR is a minor pathway for linkage analysis to determine the order of factors (genes) recombinational repair of DSBs that occur in mitotically- on a chromosome, thus simultaneously establishing that proliferating cells and that the synthesis-dependent genes are located at discrete physical locations along chromo- strand annealing (SDSA) model appears to describe somes as well as originating the classic tool of genetic map- mitotic DSB repair more accurately. -
The Role of Blm Helicase in Homologous Recombination, Gene Conversion Tract Length, and Recombination Between Diverged Sequences in Drosophila Melanogaster
| INVESTIGATION The Role of Blm Helicase in Homologous Recombination, Gene Conversion Tract Length, and Recombination Between Diverged Sequences in Drosophila melanogaster Henry A. Ertl, Daniel P. Russo, Noori Srivastava, Joseph T. Brooks, Thu N. Dao, and Jeannine R. LaRocque1 Department of Human Science, Georgetown University Medical Center, Washington, DC 20057 ABSTRACT DNA double-strand breaks (DSBs) are a particularly deleterious class of DNA damage that threatens genome integrity. DSBs are repaired by three pathways: nonhomologous-end joining (NHEJ), homologous recombination (HR), and single-strand annealing (SSA). Drosophila melanogaster Blm (DmBlm) is the ortholog of Saccharomyces cerevisiae SGS1 and human BLM, and has been shown to suppress crossovers in mitotic cells and repair mitotic DNA gaps via HR. To further elucidate the role of DmBlm in repair of a simple DSB, and in particular recombination mechanisms, we utilized the Direct Repeat of white (DR-white) and Direct Repeat of white with mutations (DR-white.mu) repair assays in multiple mutant allele backgrounds. DmBlm null and helicase-dead mutants both demonstrated a decrease in repair by noncrossover HR, and a concurrent increase in non-HR events, possibly including SSA, crossovers, deletions, and NHEJ, although detectable processing of the ends was not significantly impacted. Interestingly, gene conversion tract lengths of HR repair events were substantially shorter in DmBlm null but not helicase-dead mutants, compared to heterozygote controls. Using DR-white.mu,we found that, in contrast to Sgs1, DmBlm is not required for suppression of recombination between diverged sequences. Taken together, our data suggest that DmBlm helicase function plays a role in HR, and the steps that contribute to determining gene conversion tract length are helicase-independent. -
A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated. -
ATRX Protects Cells Against Replication-Induced Genomic Instability
ATRX Protects Cells Against Replication-Induced Genomic Instability Danton Ivanochko M.Sc. Thesis This thesis is submitted to the Faculty of Graduate and Postdoctoral Studies in partial fulfillment of the requirements for the degree of Master of Science in Biochemistry with a specialization in Human and Molecular Genetics Department of Biochemistry, Microbiology, and Immunology Faculty of Medicine University of Ottawa, Ontario, Ontario, Canada © Danton Ivanochko, Ottawa, Canada, 2016 Dedicated to my parents. ii Abstract Expansive proliferation of neural progenitor cells (NPCs) is a prerequisite to the temporal waves of neuronal differentiation that generate the six-layered cerebral cortex. NPC expansion places a heavy burden on proteins that regulate chromatin packaging and genome integrity, which is further reflected by the growing number of developmental disorders caused by mutations in chromatin regulators. Accordingly, mutations in ATRX, a chromatin remodelling protein required for heterochromatin maintenance at telomeres and simple repeats, cause the ATR-X syndrome. Here, we demonstrate that proliferating ATRX-null cells accumulate DNA damage, while also exhibiting sensitivity to hydroxyurea-induced replication fork stalling. Specifically, PARP1 hyperactivation and replication-dependent double strand DNA breakage indicated replication fork protection defects, while DNA fiber assays confirmed that ATRX was required to protect replication forks from degradation. Interestingly, inhibition of the exonuclease MRE11 by the small molecule mirin could prevent degradation. Thus, ATRX is required to limit replication stress during NPC expansion. iii Acknowledgements First and foremost, I would like to thank my supervisor, Dr. David Picketts, for his guidance and support during my undergraduate and graduate studies. Thanks to the members of my thesis advisory committee, Dr. -
000466 SIMR REPRT Fall2k3
NEWS AND THE INSIGHT FROM THE STOWERS INSTITUTE FOR MEDICAL Stowers RESEARCH REPORT FALL 2 0 0 Stowers Institute for Medical Research principal investigators who have received recent noteworthy awards and honors gather at the west end 3 of the Stowers Institute® campus. Front row, from left: Paul Trainor, Robb Krumlauf, Chunying Du. Second row, from left: Olivier Pourquié, Peter Baumann, Jennifer Gerton, Ting Xie. Ultimate solutions take time. Inside this issue . That’s particularly true with complex • Dr. Scott Hawley makes some surprising discoveries about how mistakes human diseases and birth defects during meiosis can lead to miscarriages and birth defects (Page 2). since there is still much we don’t • Dr. Olivier Pourquié sheds light on how the segments of the body begin to understand about the fundamentals grow at the right time and place in the embryo (Page 4). of life. At the Stowers Institute for • How do cells know when and where to differentiate and when their useful Medical Research, investigators healthy life is over? Dr. Chunying Du discovers a curious double negative seek to increase the understanding feedback loop in the apoptosis process that goes awry in cancer (Page 6); of the basic processes in living cells – Dr. Ting Xie investigates the importance of an environmental niche for VOLUME 6 stem cells (Page 7); and Dr. Peter Baumann studies the role of telomeres in a crucial step in the search for new aging and cancer (Page 8). medical treatments. • Scientific Director Dr. Robb Krumlauf and fellow Stowers Institute investigators inspire and are inspired by scientists and students in embryology at the Marine Biological Laboratory in Woods Hole, Massachusetts (Page 10). -
Mechanisms and Regulation of Mitotic Recombination in Saccharomyces Cerevisiae
YEASTBOOK GENOME ORGANIZATION AND INTEGRITY Mechanisms and Regulation of Mitotic Recombination in Saccharomyces cerevisiae Lorraine S. Symington,* Rodney Rothstein,† and Michael Lisby‡ *Department of Microbiology and Immunology, and yDepartment of Genetics and Development, Columbia University Medical Center, New York, New York 10032, and ‡Department of Biology, University of Copenhagen, DK-2200 Copenhagen, Denmark ABSTRACT Homology-dependent exchange of genetic information between DNA molecules has a profound impact on the maintenance of genome integrity by facilitating error-free DNA repair, replication, and chromosome segregation during cell division as well as programmed cell developmental events. This chapter will focus on homologous mitotic recombination in budding yeast Saccharomyces cerevisiae.However, there is an important link between mitotic and meiotic recombination (covered in the forthcoming chapter by Hunter et al. 2015) and many of the functions are evolutionarily conserved. Here we will discuss several models that have been proposed to explain the mechanism of mitotic recombination, the genes and proteins involved in various pathways, the genetic and physical assays used to discover and study these genes, and the roles of many of these proteins inside the cell. TABLE OF CONTENTS Abstract 795 I. Introduction 796 II. Mechanisms of Recombination 798 A. Models for DSB-initiated homologous recombination 798 DSB repair and synthesis-dependent strand annealing models 798 Break-induced replication 798 Single-strand annealing and microhomology-mediated end joining 799 B. Proteins involved in homologous recombination 800 DNA end resection 800 Homologous pairing and strand invasion 802 Rad51 mediators 803 Single-strand annealing 803 DNA translocases 804 DNA synthesis during HR 805 Resolution of recombination intermediates 805 III. -
Chromosome Segregation: Learning Only When Chromosomes Are Correctly Bi-Oriented and Microtubules Exert to Let Go Tension Across Sister Kinetochores [6]
View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector Dispatch R883 an mTORC1 substrate that negatively regulates inhibitors. Oncogene http://dx.doi.org/10.1038/ 1Department of Cancer and Cell Biology, insulin signaling. Science 332, 1322–1326. onc.2013.92. University of Cincinnati College of Medicine, 16. Chung, J., Kuo, C.J., Crabtree, G.R., and 19. She, Q.B., Halilovic, E., Ye, Q., Zhen, W., Cincinnati, OH 45267, USA. 2Institute for Blenis, J. (1992). Rapamycin-FKBP specifically Shirasawa, S., Sasazuki, T., Solit, D.B., and blocks growth-dependent activation of and Rosen, N. (2010). 4E-BP1 is a key effector of the Research in Immunology and Cancer (IRIC), signaling by the 70 kd S6 protein kinases. Cell oncogenic activation of the AKT and ERK Universite´ de Montre´ al, Montreal, 69, 1227–1236. signaling pathways that integrates their Quebec H3C 3J7, Canada. 3Department of 17. Zhang, Y., and Zheng, X.F. (2012). function in tumors. Cancer Cell 18, Pathology and Cell Biology, Faculty of mTOR-independent 4E-BP1 phosphorylation is 39–51. Medicine, Universite´ de Montre´ al, Montreal, associated with cancer resistance to mTOR 20. Shin, S., Wolgamott, L., Tcherkezian, J., kinase inhibitors. Cell Cycle 11, 594–603. Vallabhapurapu, S., Yu, Y., Roux, P.P., and Quebec, H3C 3J7, Canada. 18. Ducker, G.S., Atreya, C.E., Simko, J.P., Yoon, S.O. (2013). Glycogen synthase E-mail: [email protected], philippe. Hom, Y.K., Matli, M.R., Benes, C.H., Hann, B., kinase-3beta positively regulates protein [email protected] Nakakura, E.K., Bergsland, E.K., Donner, D.B., synthesis and cell proliferation through the et al. -
Accurate Chromosome Segregation by Probabilistic Self-Organisation Yasushi Saka1*, Claudiu V
Saka et al. BMC Biology (2015) 13:65 DOI 10.1186/s12915-015-0172-y RESEARCH ARTICLE Open Access Accurate chromosome segregation by probabilistic self-organisation Yasushi Saka1*, Claudiu V. Giuraniuc1 and Hiroyuki Ohkura2* Abstract Background: For faithful chromosome segregation during cell division, correct attachments must be established between sister chromosomes and microtubules from opposite spindle poles through kinetochores (chromosome bi-orientation). Incorrect attachments of kinetochore microtubules (kMTs) lead to chromosome mis-segregation and aneuploidy, which is often associated with developmental abnormalities such as Down syndrome and diseases including cancer. The interaction between kinetochores and microtubules is highly dynamic with frequent attachments and detachments. However, it remains unclear how chromosome bi-orientation is achieved with such accuracy in such a dynamic process. Results: To gain new insight into this essential process, we have developed a simple mathematical model of kinetochore–microtubule interactions during cell division in general, i.e. both mitosis and meiosis. Firstly, the model reveals that the balance between attachment and detachment probabilities of kMTs is crucial for correct chromosome bi-orientation. With the right balance, incorrect attachments are resolved spontaneously into correct bi-oriented conformations while an imbalance leads to persistent errors. In addition, the model explains why errors are more commonly found in the first meiotic division (meiosis I) than in mitosis and how a faulty conformation can evade the spindle assembly checkpoint, which may lead to a chromosome loss. Conclusions: The proposed model, despite its simplicity, helps us understand one of the primary causes of chromosomal instability—aberrant kinetochore–microtubule interactions. The model reveals that chromosome bi-orientation is a probabilistic self-organisation, rather than a sophisticated process of error detection and correction. -
Patient Mutations Alter ATRX Targeting to PML Nuclear Bodies
European Journal of Human Genetics (2008) 16, 192–201 & 2008 Nature Publishing Group All rights reserved 1018-4813/08 $30.00 www.nature.com/ejhg ARTICLE Patient mutations alter ATRX targeting to PML nuclear bodies Nathalie G Be´rube´1,3,4, Jasmine Healy1,4, Chantal F Medina1, Shaobo Wu1, Todd Hodgson1, Magdalena Jagla1 and David J Picketts*,2 1Molecular Medicine Program, Ottawa Health Research Institute, Ottawa, Ontario, Canada; 2Departments of Medicine and Biochemistry, Microbiology, and Immunology, University of Ottawa, Ottawa, Ontario, Canada ATRX is a SWI/SNF-like chromatin remodeling protein mutated in several X-linked mental retardation syndromes. Gene inactivation studies in mice demonstrate that ATRX is an essential protein and suggest that patient mutations likely retain partial activity. ATRX associates with the nuclear matrix, pericentromeric heterochromatin, and promyelocytic leukemia nuclear bodies (PML-NBs) in a speckled nuclear staining pattern. Here, we used GFP–ATRX fusion proteins to identify the specific domains of ATRX necessary for subnuclear targeting and the effect of patient mutations on this localization. We identified two functional nuclear localization signals (NLSs) and two domains that target ATRX to nuclear speckles. One of the latter domains is responsible for targeting ATRX to PML-NBs. Surprisingly, this domain encompassed motifs IV–VI of the SNF2 domain suggesting that in addition to chromatin remodeling, it may also have a role in subnuclear targeting. More importantly, four different patient mutations within this domain resulted in an B80% reduction in the number of transfected cells with ATRX nuclear speckles and PML colocalization. These results demonstrate that patient mutations have a dramatic effect on subnuclear targeting to PML-NBs. -
X-Linked Diseases: Susceptible Females
REVIEW ARTICLE X-linked diseases: susceptible females Barbara R. Migeon, MD 1 The role of X-inactivation is often ignored as a prime cause of sex data include reasons why women are often protected from the differences in disease. Yet, the way males and females express their deleterious variants carried on their X chromosome, and the factors X-linked genes has a major role in the dissimilar phenotypes that that render women susceptible in some instances. underlie many rare and common disorders, such as intellectual deficiency, epilepsy, congenital abnormalities, and diseases of the Genetics in Medicine (2020) 22:1156–1174; https://doi.org/10.1038/s41436- heart, blood, skin, muscle, and bones. Summarized here are many 020-0779-4 examples of the different presentations in males and females. Other INTRODUCTION SEX DIFFERENCES ARE DUE TO X-INACTIVATION Sex differences in human disease are usually attributed to The sex differences in the effect of X-linked pathologic variants sex specific life experiences, and sex hormones that is due to our method of X chromosome dosage compensation, influence the function of susceptible genes throughout the called X-inactivation;9 humans and most placental mammals – genome.1 5 Such factors do account for some dissimilarities. compensate for the sex difference in number of X chromosomes However, a major cause of sex-determined expression of (that is, XX females versus XY males) by transcribing only one disease has to do with differences in how males and females of the two female X chromosomes. X-inactivation silences all X transcribe their gene-rich human X chromosomes, which is chromosomes but one; therefore, both males and females have a often underappreciated as a cause of sex differences in single active X.10,11 disease.6 Males are the usual ones affected by X-linked For 46 XY males, that X is the only one they have; it always pathogenic variants.6 Females are biologically superior; a comes from their mother, as fathers contribute their Y female usually has no disease, or much less severe disease chromosome. -
A Yeast Phenomic Model for the Influence of Warburg Metabolism on Genetic Buffering of Doxorubicin Sean M
Santos and Hartman Cancer & Metabolism (2019) 7:9 https://doi.org/10.1186/s40170-019-0201-3 RESEARCH Open Access A yeast phenomic model for the influence of Warburg metabolism on genetic buffering of doxorubicin Sean M. Santos and John L. Hartman IV* Abstract Background: The influence of the Warburg phenomenon on chemotherapy response is unknown. Saccharomyces cerevisiae mimics the Warburg effect, repressing respiration in the presence of adequate glucose. Yeast phenomic experiments were conducted to assess potential influences of Warburg metabolism on gene-drug interaction underlying the cellular response to doxorubicin. Homologous genes from yeast phenomic and cancer pharmacogenomics data were analyzed to infer evolutionary conservation of gene-drug interaction and predict therapeutic relevance. Methods: Cell proliferation phenotypes (CPPs) of the yeast gene knockout/knockdown library were measured by quantitative high-throughput cell array phenotyping (Q-HTCP), treating with escalating doxorubicin concentrations under conditions of respiratory or glycolytic metabolism. Doxorubicin-gene interaction was quantified by departure of CPPs observed for the doxorubicin-treated mutant strain from that expected based on an interaction model. Recursive expectation-maximization clustering (REMc) and Gene Ontology (GO)-based analyses of interactions identified functional biological modules that differentially buffer or promote doxorubicin cytotoxicity with respect to Warburg metabolism. Yeast phenomic and cancer pharmacogenomics data were integrated to predict differential gene expression causally influencing doxorubicin anti-tumor efficacy. Results: Yeast compromised for genes functioning in chromatin organization, and several other cellular processes are more resistant to doxorubicin under glycolytic conditions. Thus, the Warburg transition appears to alleviate requirements for cellular functions that buffer doxorubicin cytotoxicity in a respiratory context. -
Branching Out: Meiotic Recombination and Its Regulation
TICB-453; No of Pages 8 Review TRENDS in Cell Biology Vol.xxx No.x Branching out: meiotic recombination and its regulation Gareth A. Cromie and Gerald R. Smith Division of Basic Sciences, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue North, Seattle, WA 98109-1024, USA Homologous recombination is a dynamic process by parental chromosome segregation during the first meiotic which DNA sequences and strands are exchanged. In division. The COs link the homologous chromosomes phy- meiosis, the reciprocal DNA recombination events called sically so that they can be oriented correctly on the meiotic crossovers are central to the generation of genetic diver- spindle. In the absence of COs, chromosomes often mis- sity in gametes and are required for homolog segregation segregate, resulting in aneuploid gametes and offspring. in most organisms. Recent studies have shed light on how Recent studies have advanced our understanding of how meiotic crossovers and other recombination products meiotic COs and NCOs form, how they are distributed form, how their position and number are regulated and across genomes, and how the pair of DNA molecules under- how the DNA molecules undergoing recombination are going a CO is chosen. In this review, we focus on how chosen. These studies indicate that the long-dominant, advances in these three areas have challenged several core unifying model of recombination proposed by Szostak features of long-accepted models, revealing many new et al. applies, with modification, only to a subset of branches of the meiotic recombination ‘pathway’. Most recombination events. Instead, crossover formation and significantly, the mechanism of recombination associated its control involve multiple pathways, with considerable with the well-known DSB repair model of Szostak et al.