Regulation of DNA double-strand break repair pathway choice
Regulation of DNA double-strand break repair pathway choice
批准号:
10656805
负责人:
Wolf-Dietrich Heyer
金额:
$46.66万
依托单位国家:
美国
项目类别:
财政年份:
2023
资助国家:
美国
项目状态:
未结题
起止时间:
2023-07-01 至 2028-06-30
关键词:
AbbreviationsAffectBRCA2 geneBase PairingBiochemicalBiochemical ReactionC-terminalCamptothecinCell CycleCellsChromosomal RearrangementClinicalComplexCyclin-Dependent KinasesDNA BindingDNA Binding DomainDNA DamageDNA Double Strand BreakDNA Repair GeneDNA biosynthesisDNA damage checkpointDNA lesionDNA replication forkDNA-Directed DNA PolymeraseDataDefectDouble Strand Break RepairDrug TargetingEnergy TransferFilamentFluorescenceFluorescence Resonance Energy TransferGenesGenomeGenome StabilityGenomicsGoalsHumanImageImmune checkpoint inhibitorIn VitroIndividualIonizing radiationKnowledgeLengthMediatingMediatorMitosisModalityModelingMutagenesisNonhomologous DNA End JoiningPathway interactionsPatient SelectionPhasePoly(ADP-ribose) Polymerase InhibitorPoly(ADP-ribose) PolymerasesPolymerasePredispositionProcessProductivityPropertyProteinsProtocols documentationRAD52 geneRad51 recombinaseRadiation induced damageReactionReactive Oxygen SpeciesRegimenRegulationResearchResistanceRoleS phaseSideSourceTestingTherapeuticTopoisomerase IIType I DNA Topoisomerasescancer cellcancer therapychromosome fusionchromosome lossclinical applicationhomologous recombinationinhibitorinnovationinsightloss of functionmutantnovelpersonalized medicineprotein purificationreconstitutionrepairedresponsetranslational potentialtumor
中文摘要
项目总结
英文摘要
Project Summary
The general goal of the proposed research is to define the mechanistic underpinning for the synthetic lethality
network between a deficiency in homologous recombination (HR) and defects in single-strand annealing
(RAD52) or POLq-mediated end-joining (TMEJ). Exciting preliminary data revealed that BRCA2 and RAD52
delay the repair of S-phase-associated DNA double-stranded breaks (DSB) by TMEJ until M-phase. This
regulation avoids TMEJ-mediated chromosomal rearrangements of one-sided DSBs that are produced by
replication fork breakage. Our approach combines innovative cell cycle phase resolved imaging of DNA repair
proteins and DNA damage markers with mechanistic biochemical analysis using purified human proteins in
reconstituted reactions. Our results will have potential translational implications for the clinical application of
newly developed RAD52 and POLq inhibitors for the treatment of HR-deficient tumors with respect to application
protocols, patient selection, and use of DNA damage response checkpoint inhibitors as well as the response to
poly(ADP-ribose) polymerase inhibition.
The Specific Aims are:
1. Define the mechanism of action of BRCA2 in DSB repair pathway control. We will test the model that the
DNA binding properties of BRCA2 are critical for TMEJ inhibition. In Aim 1A, we conduct foundational studies
to determine the fundamental DNA binding properties of full-length BRCA2. In Aim 1B, we will define which
domains of BRCA2 are required for TMEJ inhibition in cells. This combination of cell-based and biochemical
studies will define the functions and regions of BRCA2 that are required for TMEJ inhibition.
2. Define the mechanism of TMEJ inhibition by BRCA2 and RAD52. BRCA2 and RAD52 employ two different
modes to inhibit the DNA polymerase activity of POLq which may affect additional reaction steps in the TMEJ
process. We will reconstitute TMEJ in vitro with purified proteins to determine the mechanisms by which
BRCA2 (Aim 2A) and RAD52 (Aim 2B) inhibit TMEJ. We will test inhibition of the overall TMEJ reaction and
individual steps including 1) DNA binding, 2) end-alignment, and 3) DNA synthesis. Analysis of wild type and
catalytic mutants of POLq will be conducted in vitro and in cells.
3. Define which HR defects are susceptible to RAD52 loss of function. It is an open question whether loss
of RAD52 will lead to POLq-mediated chromosome fusions and lethality in all HR-deficient backgrounds (Aim
3A) or all BRCA2 mutants (Aim 3B). Our preliminary studies suggest a model that loading of BRCA2 is the
critical step to limit TMEJ to M-phase and that HR defects past this step are not affected by RAD52 inhibition.
期刊论文(0)
专著(0)
科研奖励(0)
会议论文
Genome instability induced by homologous recombination
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批准号:10019104
-
项目类别:
-
资助金额:$31.4万
-
财政年份:2020
-
负责人:Wolf-Dietrich Heyer
-
依托单位:
Genome instability induced by homologous recombination
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批准号:10437930
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项目类别:
-
资助金额:$31.4万
-
财政年份:2020
-
负责人:Wolf-Dietrich Heyer
-
依托单位:
Genome instability induced by homologous recombination
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批准号:10795314
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项目类别:
-
资助金额:$1.92万
-
财政年份:2020
-
负责人:Wolf-Dietrich Heyer
-
依托单位:
Genome instability induced by homologous recombination
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批准号:10241424
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项目类别:
-
资助金额:$31.4万
-
财政年份:2020
-
负责人:Wolf-Dietrich Heyer
-
依托单位:
Genome instability induced by homologous recombination
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批准号:10670239
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项目类别:
-
资助金额:$31.4万
-
财政年份:2020
-
负责人:Wolf-Dietrich Heyer
-
依托单位:
Genome instability induced by homologous recombination
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批准号:10806624
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项目类别:
-
资助金额:$1.13万
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财政年份:2020
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负责人:Wolf-Dietrich Heyer
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依托单位:
2015 Nucleic Acids Gordon Research Conference
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批准号:8892554
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项目类别:
-
资助金额:$0.55万
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财政年份:2015
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负责人:Wolf-Dietrich Heyer
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依托单位:
FASEB SRC: Genetic Recombination and Genome Rearrangements
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批准号:8525768
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项目类别:
-
资助金额:$0.3万
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财政年份:2013
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负责人:Wolf-Dietrich Heyer
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依托单位:
Functions of Rad51 Paralogs in Recombinational DNA Repair
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批准号:8519965
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项目类别:
-
资助金额:$48.87万
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财政年份:2011
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负责人:Wolf-Dietrich Heyer
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依托单位:
Functions of Rad51 Paralogs in Recombinational DNA Repair
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批准号:8890651
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项目类别:
-
资助金额:$53.03万
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财政年份:2011
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负责人:Wolf-Dietrich Heyer
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依托单位:
Functions of Rad51 Paralogs in Recombinational DNA Repair
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批准号:8703631
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项目类别:
-
资助金额:$51.44万
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财政年份:2011
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负责人:Wolf-Dietrich Heyer
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依托单位:
Functions of Rad51 Paralogs in Recombinational DNA Repair
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批准号:8338889
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项目类别:
-
资助金额:$51.99万
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财政年份:2011
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负责人:Wolf-Dietrich Heyer
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依托单位:
Functions of Rad51 Paralogs in Recombinational DNA Repair
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批准号:8237897
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项目类别:
-
资助金额:$51.83万
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财政年份:2011
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负责人:Wolf-Dietrich Heyer
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依托单位:
DNA DAMAGE CHECKPOINT MEDIATED REGULATION OF DOUBLE-STRAND BREAK REPAIR
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批准号:8171262
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项目类别:
-
资助金额:$0.08万
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财政年份:2010
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负责人:Wolf-Dietrich Heyer
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依托单位:
RECOMBINATIONAL MECHANISMS OF DNA REPAIR IN EUKARYOTES
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批准号:8171388
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项目类别:
-
资助金额:$0.24万
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财政年份:2010
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负责人:Wolf-Dietrich Heyer
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依托单位:
DNA DAMAGE CHECKPOINT MEDIATED REGULATION OF DOUBLE-STRAND BREAK REPAIR
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批准号:7957786
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项目类别:
-
资助金额:$0.33万
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财政年份:2009
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负责人:Wolf-Dietrich Heyer
-
依托单位:
RECOMBINATIONAL MECHANISMS OF DNA REPAIR IN EUKARYOTES
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批准号:7957775
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项目类别:
-
资助金额:$0.33万
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财政年份:2009
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负责人:Wolf-Dietrich Heyer
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依托单位:
RECOMBINATIONAL MECHANISMS OF DNA REPAIR IN EUKARYOTES
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批准号:7723651
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项目类别:
-
资助金额:$0.81万
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财政年份:2008
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负责人:Wolf-Dietrich Heyer
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依托单位:
Microbial Production Core Facility
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批准号:7211306
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项目类别:
-
资助金额:$22.64万
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财政年份:2007
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负责人:Wolf-Dietrich Heyer
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依托单位:
DNA DAMAGE CHECKPOINT MEDIATED REGULATION OF DOUBLE-STRAND BREAK REPAIR
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批准号:7602190
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项目类别:
-
资助金额:$0.08万
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财政年份:2007
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负责人:Wolf-Dietrich Heyer
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依托单位:
海外基金