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Mapping repair-relevant domains of the nuclear envelope by superresolution microscopy

English title Mapping repair-relevant domains of the nuclear envelope by superresolution microscopy
Applicant Gasser Susan
Number 141945
Funding scheme Sinergia
Research institution Friedrich Miescher Institute for Biomedical Research
Institution of higher education Institute Friedrich Miescher - FMI
Main discipline Cellular Biology, Cytology
Start/End 01.08.2012 - 31.03.2016
Approved amount 1'200'000.00
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All Disciplines (2)

Discipline
Cellular Biology, Cytology
Biophysics

Keywords (9)

DNA repair; Nuclear envelope; Yeast; Superresolution microscopy; Chemistry; single molecule localization; Bicyclic compounds; Telomeres; C elegans

Lay Summary (English)

Lead
Lay summary

In this project a synthetic chemist, a geneticist and a physicist with microscopy expertise will use novel tagging methods and super-resolution microscopy to examine the proteins that create functional subdomains on the inner face of the nuclear envelope.  The cell nucleus harbors all the cell’s genetic information and coordinates its conversion into messenger RNA, its replication, repair and segregation.  These are essential functions of life. The jumble of DNA is organized into functional domains, in large part through interaction with structures at the nuclear surface – or more precisely on the inner face of a double envelope that surrounds the nucleus.  To understand the organization of the genome through its interaction sites,  we must develop novel imaging tools – both superresolution microscopy methods that go beyond the canonical resolution limits, and small but specific ligands that will distinguish one type of molecule from another. This project aims to combine such technologies with powerful genetic models of yeast and C elegans.  The result should explain better how genome interaction with the nuclear envelope controls its function. Several human diseases are correlated with loss of genomic integrity due to alterations of nuclear envelope- genome interactions, and the results of this study will provide tools to explore the causal relationships implied by this disease correlation. 

Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Asymmetric Processing of DNA Ends at a Double-Strand Break Leads to Unconstrained Dynamics and Ectopic Translocation
MarcominiIsabella, ShimadaKenji, DelgoshaieNeda, Yamamoto Io, SeeberAndrew, Cheblalanais, HorigomeChihiro, NaumannUlrike, GasserSusan (2018), Asymmetric Processing of DNA Ends at a Double-Strand Break Leads to Unconstrained Dynamics and Ectopic Translocation, in Cell Reports, 24 (10), 2614-2628.
Histones and histone modifications in perinuclear chromatin anchoring: from yeast to man
Harr JC, Gonzalez-Sandoval A, Gasser SM (2016), Histones and histone modifications in perinuclear chromatin anchoring: from yeast to man, in EMBO Report, 2, 139-155.
PolySUMOylation by Siz2 and Mms21 triggers relocation of DNA breaks to nuclear pores through the Slx5/Slx8 STUbL
Horigiome C, Bustard DE, Marcomini I, Delgoshaie N, Tsai-Pflugfelder M, Cobb JA, Gasser SM (2016), PolySUMOylation by Siz2 and Mms21 triggers relocation of DNA breaks to nuclear pores through the Slx5/Slx8 STUbL, in Genes Develpment, 30(8), 931-945.
Spatial Control Over Gene Expression
Gonzalez-Sandoval A, Gasser SM (2016), Spatial Control Over Gene Expression, in Trends Genetic, June 13, 1.
A simple method for GFP- and RFP-based dual color single-molecule localization microscopy
Platonova E, Winterflood CM, Ewers H (2015), A simple method for GFP- and RFP-based dual color single-molecule localization microscopy, in ACS Chem Biol, 10(6), 1411-1416.
Absolute Arrangement of Subunits in Cytoskeletal Septin Filaments in Cells Measured by Fluorescence Microscopy
Kaplan C, Jing B, Winterflood CM, Bridges AA, Occhipinti P, Schmid J, Grinhagens S, Gronemeyer T, Tinnefeld P, Ewers H (2015), Absolute Arrangement of Subunits in Cytoskeletal Septin Filaments in Cells Measured by Fluorescence Microscopy, in Nano Lett, 15(6), 3859-3854.
Dual-color 3D superresolution microscopy by combined spectral-demixing and biplane imaging
Winterflood CM, Platonova E, Albrecht D, Ewers H (2015), Dual-color 3D superresolution microscopy by combined spectral-demixing and biplane imaging, in Biophys J, 109(1), 3-6.
Nuclear organization in DNA end processing: Telomeres vs double-strand breaks
Marcomini I, Gasser SM (2015), Nuclear organization in DNA end processing: Telomeres vs double-strand breaks, in DNA Repair (Amst), 32, 134-140.
Optimized sample preparation for single-molecule localization-based superresolution microscopy in yeast
Kaplan C, Ewers H (2015), Optimized sample preparation for single-molecule localization-based superresolution microscopy in yeast, in Nat Protoc, 10(7), 1007-1021.
Perinuclear Anchoring of H3K9-Methylated Chromatin Stabilizes Induced Cell Fate in C. elegans Embryos
Gonzalez-Sandoval A, Towbin BD, Kalck V, Cabianca DS, Gaidatzis D, Hauer MH, Geng L, Wang L, Yang T, Gasser SM (2015), Perinuclear Anchoring of H3K9-Methylated Chromatin Stabilizes Induced Cell Fate in C. elegans Embryos, in Cell, 163(6), 1333-1347.
Regulation of recombination atyeast nuclear pores controls repair and triplet repeat stability
Su XA, Dion V, Gasser SM, Freundenreich CH (2015), Regulation of recombination atyeast nuclear pores controls repair and triplet repeat stability, in Genes Development, 29(10), 1006-1017.
Resolving bundled microtubules using anti-tubulin nanobodies
Mikhaylova M, Cloin BM, Finan K, van den Berg R, Teeuw J, Kijanka MM, Sokolowski M, Katrukha EA, Miadorn M, Kapitein LC (2015), Resolving bundled microtubules using anti-tubulin nanobodies, in Nat Commun, 6, 7933.
Single-molecule microscopy of molecules tagged with GFP or RFP derivatives in mammalian cells using nanobody binders
Platonova E, Winterflood CM, Junemann A, Albrecht D, Faix J, Ewers H (2015), Single-molecule microscopy of molecules tagged with GFP or RFP derivatives in mammalian cells using nanobody binders, in Methods, 88, 89-97.
Visualizing the spatiotemporal dynamics of DNA damage in budding yeast
Horigome C, Dion V, Seeber A, Gehlen LR, Gasser SM (2015), Visualizing the spatiotemporal dynamics of DNA damage in budding yeast, in Methods Mol Biol., 1292, 77-96.
Drug discovery: Tools and rules for macrocycles
Heinis C (2014), Drug discovery: Tools and rules for macrocycles, in Nat Chem Biol, Jul 20, 1.
Peptide ligands stabilized by small molecules
Chen S, Bertoldo D, Angelini A, Pojer F, Heinis C (2014), Peptide ligands stabilized by small molecules, in Angew Chem Int Ed Engl., 53(6), 1602-1606.
Phage selection of photoswitchable peptide ligands
Bellotto S, Chen S, Rebollo I, Wegner HA, Heinis C (2014), Phage selection of photoswitchable peptide ligands, in J Am Chem Soc, 136(16), 5880-5883.
Single-molecule localization microscopy using mCherry
Winterflood CM, Ewers H (2014), Single-molecule localization microscopy using mCherry, in Chemphyschem, 15(16), 3447-3451.
Tracking chemical reactions on the surface of filamentous phage using mass spectrometry
Chen S, Touati J, Heinis C (2014), Tracking chemical reactions on the surface of filamentous phage using mass spectrometry, in Chem Commun (Camb), 50(40), 5267-5269.
Promoter- and RNA polymerase II-dependent hsp-16 gene association with nuclear pores in Caenorhabditis elegans
Rohner S, Kalck V, Wang X, Ikegami K, Lieb JD, Gasser SM, Meister P (2013), Promoter- and RNA polymerase II-dependent hsp-16 gene association with nuclear pores in Caenorhabditis elegans, in J. Cell Biol., 200(5), 589-604.
Chromatin movement in the maintenance of genome stability
Dion V. and Gasser S.M. (2013), Chromatin movement in the maintenance of genome stability, in Cell, 152, 1255 -1364.
Improving binding affinity and stability of peptide ligands by substituting glycines with D-amino acids
Chen S, Gfeller D, Buth SA, Michielin O, Leiman PG, Heinis C (2013), Improving binding affinity and stability of peptide ligands by substituting glycines with D-amino acids, in Chembiochem, 14(11), 1316-1322.
Superresolution imaging of amyloid fibrils with binding-activated probes
Ries J, Udayar V, Soragni A, Hornemann S, Nilsson KP, Riek R, Hock C, Ewers H, Aguzzi AA, Rajendran L (2013), Superresolution imaging of amyloid fibrils with binding-activated probes, in ACS Chem Neurosci, 4(7), 1057-1061.
The bacterial SMC complex displays two distinct modes of interaction with the chromosome
Kleine Borgmann LA, Ries J, Ewers H, Ulbrich MH, Graumann PL (2013), The bacterial SMC complex displays two distinct modes of interaction with the chromosome, in Cell Rep, 3(5), 1483-1492.
The shelterin protein POT-1 anchors Caenorhabditis elegans telomeres through SUN-1 at the nuclear periphery
Ferreira HC, Towbin BD, Jegou T, Gasser SM (2013), The shelterin protein POT-1 anchors Caenorhabditis elegans telomeres through SUN-1 at the nuclear periphery, in J. Cell. Biol, 203(5), 727-735.
The shelterin protein POT-1 anchors Caenorhabditis elegans telomeres through SUN-1 at the nuclear periphery.
Ferreira Helder C, Towbin Benjamin D, Jegou Thibaud, Gasser Susan M (2013), The shelterin protein POT-1 anchors Caenorhabditis elegans telomeres through SUN-1 at the nuclear periphery., in The Journal of cell biology, 203(5), 727-35.
Spatial segregation of heterochromatin: uncovering functionality in a multicellular organism
Cabianca DS, Gasser SM, Spatial segregation of heterochromatin: uncovering functionality in a multicellular organism, in Nucleus, May 17, 1.
Swr1 and INO80 Chromatin Remodelers Contribute to DNA Double-Strand Break Perinuclear Anchorage Site Choice.
Horigome Chihiro, Oma Yukako, Konishi Tatsunori, Schmid Roger, Marcomini Isabella, Hauer Michael H, Dion Vincent, Harata Masahiko, Gasser Susan M, Swr1 and INO80 Chromatin Remodelers Contribute to DNA Double-Strand Break Perinuclear Anchorage Site Choice., in Molecular cell.

Collaboration

Group / person Country
Types of collaboration
M. HArata at Tohoku University, Sendai Japan (Asia)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel

Scientific events



Self-organised

Title Date Place
Development of bicyclic peptides for biological use - one day symposium 30.05.2014 FMI Basel, Switzerland

Knowledge transfer events



Self-organised

Title Date Place
FMI Annual Retreat 18.09.2013 Arosa, Switzerland

Communication with the public

Communication Title Media Place Year
New media (web, blogs, podcasts, news feeds etc.) Damage at the nuclear edge Website summary and release re MOl Cell article Western Switzerland German-speaking Switzerland International 2014

Awards

Title Year
dr honoris cause. Charles University of Prague 2016 2016
dr honoris causa: university of lausanne 2015 2015
Weizman Woman in Science award, to Susan Gasser, 2013 University of Lausanne, Doctorat honoris causa, 2014 Japanese Society for the promotion of Science postdoctoral fellowship to Chihiro Horigome (2013) Marie Curie European mobility postdoctoral fellowship (2014) 2013

Associated projects

Number Title Start Funding scheme
138334 Long-range Chromatin Organization and Genome Stability 01.01.2012 Project funding (Div. I-III)
156936 Long-range chromatin organization and genome stability 01.01.2015 Project funding (Div. I-III)
157842 Novel bioconjugation strategy and application for drug development 01.07.2015 Temporary Backup Schemes

Abstract

Spatial analysis of proteins involved in the processing and repair of chromosome ends by super-resolution microscopy, exploiting bicyclic peptidic binders in high resolution imagingThe organization of chromatin in the nucleus is one of the most basic processes that defines eukaryotic life, and the nuclear envelope is the major structural scaffold for its functional organization. The nuclear envelope is clearly compartmentalized, most molecular players remain to be identified and the exact organization is obscured by the resolution limit of light microscopy. Even present superresolution microscopy methods cannot resolve chromatin attachment points or provide the throughput required to determine, where dozens of proteins are localized in the inner nuclear membrane. We have assembled a team of three research groups with complementary expertise for a focused project on the nanoscale organization of the eukaryotic nuclear envelope. The Gasser group is a leader in 3D organization of chromatin in the nucleus. The Ewers group has developed a novel approach for high-throughput single molecule superresolution microscopy in yeast at unprecedented resolution. The Heinis group has developed a novel method to generate small molecule 'peptidic binders' against any desired protein target.We will combine these novel technologies with the aim of -1. identifying novel nuclear envelope proteins by proteomic studies on yeast and C elegans by the Gasser lab-2. we will then use this data to generate a nanoscale map of all nuclear envelope molecules by high-throughput superresolution microscopy in the Ewers laboratory.-3. we will then develop bicyclic peptidic binders against specific molecules from the above studies, and use these to study the specific localization of telomeres and DNA-double-strand breaks in relation to prominent nuclear features such as nuclear pores in response to DNA-damage.The results of this project will be: a) the generation of a novel labeling method for superresolution microscopy that through combination of organic dyes with small molecule labeling will have unprecedented resolution. b) fundamental insight into the organization of chromatin at the nuclear envelope with enormous breadth and depth of data. c) the identification of novel mechanisms in the specific localization of telomeres and DNA-double-strand breaks.Our work will generate technological leadership in a key technology, provide insight into a fundamental biological system and may yield novel targets for the therapy of human disease.
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