The transactive response DNA-binding protein-43 (TDP-43), is an hnRNP family protein, whose intracellular aggregation has been etiologically implicated in Amyotrophic Lateral Sclerosis (ALS), a motor neuron degenerative disease (affecting two per 100,000 people worldwide) and about 40% of other common neurodegenerative diseases including Alzheimer's (AD) and Parkinson's disease (PD). Primarily involved in RNA processing, TDP-43 also binds to DNA; but its DNA binding functions are not investigated. While hereditary mutations in TDP-43 have been linked to ALS, the molecular mechanism(s) of its pathology contributing to neuronal death are still unclear. The unique feature of TDP-43 pathogenesis in ALS is its nuclear clearance and simultaneous cytoplasmic aggregation in affected motor/cortical neurons. Furthermore, significant accumulation of genomic damage is observed in TDP-43-linked diseases and previous studies identified a key DNA repair protein `Ku' in TDP-43 immunocomplex from human cells; which raises the possibility of TDP-43's involvement in DNA damage repair, which has not been investigated. In support of this, our preliminary data demonstrate: (1) involvement of nuclear TDP-43 for efficient DNA double-strand break repair (DSBR) in neurons; (2) TDP-43's recruitment at the DSB sites and stable interaction with DSBR proteins; (3) marked increase in DSB accumulation, and delayed DSB repair in TDP-43-depleted neurons and their sensitization to DSB-inducing agents; and (4) strong correlation of TDP-43 nuclear clearance/functional loss with accumulation of DSBs in ALS-affected human post-mortem spinal cord tissue. Based on these, which warrant a detailed investigation into the role of TDP-43 in neuronal genome damage response, we propose that the loss of nuclear TDP-43 causes deficient repair of DSBs in neuronal genomes, leading to persistent accumulation of lethal DSBs, which promote cell death. Thus the central goal of this project is to test this novel hypothesis to establish that TDP-43 pathology-induced DNA repair deficiency is a key etiological factor in ALS/other neurodegenerative diseases. Using state-of-the-art approaches and multiple model systems, we will comprehensively test this hypothesis by pursuing the following Specific Aims:
Aim 1 : To characterize TDP-43's involvement in DSBR in neuronal genomes, and confirm that persistent genomic damage due to TDP-43 depletion causes cell death.
Aim 2 : To establish the molecular mechanism of TDP-43's function in DSBR using our novel in vitro reconstituted DSBR assay and test the impact of ALS-linked TDP-43 mutants in repair.
Aim 3 : To unravel the linkage of TDP-43 pathology (nuclear clearance) and neurodegeneration, with the genomic damage in tissues from TDP-43 (WT/mutant) transgenic mice and human ALS postmortem brains. This grant application with many innovative aspects and based on strong preliminary data represent previously unexplored area of ALS research and understanding this phenomenon could open up new avenues of therapeutic interventions.

Public Health Relevance

Amyotrophic lateral sclerosis (ALS) is a rapidly progressing, fatal motor neuron degenerative disease, for which no effective treatment is available even for slowing down the disease progression. Multiple, complex pathologies have been implicated, including the RNA/DNA binding protein TDP-43's nucleus cytoplasm redistribution in a majority of both sporadic and inherited cases; however the molecular mechanisms of its pathology and/or physiological consequence(s) leading to neuronal death/dysfunction are still unknown. The project will establish that TDP-43 is a key component of DNA double strand break (DSB) machinery in neurons of healthy individuals and its loss of nuclear functions in ALS and other TDP-43-associated neurodegenerative diseases which include about 40% of Alzheimer's and Parkinson's disease affected patients, leads to deficient DSB repair. The resulting accumulation of lethal DSBs promotes neuronal toxicity and death. Thus this innovative grant proposal, based on strong preliminary data, will examine previously unexplored area of ALS research that would lead to a major paradigm shift in our understanding of the molecular pathogenesis of not only ALS, but also other TDP-43-associated neurodegenerative diseases; and could open up new avenues of therapeutic interventions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS088645-03
Application #
9324378
Study Section
Neural Oxidative Metabolism and Death Study Section (NOMD)
Program Officer
Gubitz, Amelie
Project Start
2015-07-01
Project End
2020-06-30
Budget Start
2017-07-01
Budget End
2018-06-30
Support Year
3
Fiscal Year
2017
Total Cost
Indirect Cost
Name
Methodist Hospital Research Institute
Department
Type
DUNS #
185641052
City
Houston
State
TX
Country
United States
Zip Code
77030
Okonkwo, Adaobi; Mitra, Joy; Johnson, Gavin S et al. (2018) Heterocyclic Analogs of Sulforaphane Trigger DNA Damage and Impede DNA Repair in Colon Cancer Cells: Interplay of HATs and HDACs. Mol Nutr Food Res 62:e1800228
Sengupta, Shiladitya; Yang, Chunying; Hegde, Muralidhar L et al. (2018) Acetylation of oxidized base repair-initiating NEIL1 DNA glycosylase required for chromatin-bound repair complex formation in the human genome increases cellular resistance to oxidative stress. DNA Repair (Amst) 66-67:1-10
Vasquez, Velmarini; Mitra, Joy; Perry, George et al. (2018) An Inducible Alpha-Synuclein Expressing Neuronal Cell Line Model for Parkinson's Disease1. J Alzheimers Dis 66:453-460
Zhang, Dan; Dong, Yiping; Zhao, Ying et al. (2018) Sinomenine hydrochloride sensitizes cervical cancer cells to ionizing radiation by impairing DNA damage response. Oncol Rep 40:2886-2895
Hegde, Muralidhar L; Bohr, Vilhelm A; Mitra, Sankar (2017) DNA damage responses in central nervous system and age-associated neurodegeneration. Mech Ageing Dev 161:1-3
Yang, Chunying; Sengupta, Shiladitya; Hegde, Pavana M et al. (2017) Regulation of oxidized base damage repair by chromatin assembly factor 1 subunit A. Nucleic Acids Res 45:739-748
Nookala, Anantha Ram; Mitra, Joy; Chaudhari, Nitish S et al. (2017) An Overview of Human Immunodeficiency Virus Type 1-Associated Common Neurological Complications: Does Aging Pose a Challenge? J Alzheimers Dis 60:S169-S193
Wang, Haibo; Dharmalingam, Prakash; Vasquez, Velmarini et al. (2017) Chronic oxidative damage together with genome repair deficiency in the neurons is a double whammy for neurodegeneration: Is damage response signaling a potential therapeutic target? Mech Ageing Dev 161:163-176
Wang, Haibo; Peng, Bin; Pandita, Raj K et al. (2017) Aurora kinase B dependent phosphorylation of 53BP1 is required for resolving merotelic kinetochore-microtubule attachment errors during mitosis. Oncotarget 8:48671-48687
German, Peter; Saenz, David; Szaniszlo, Peter et al. (2017) 8-Oxoguanine DNA glycosylase1-driven DNA repair-A paradoxical role in lung aging. Mech Ageing Dev 161:51-65

Showing the most recent 10 out of 24 publications