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1.
Elife ; 82019 04 01.
Article in English | MEDLINE | ID: mdl-30924768

ABSTRACT

While genomic sequencing routinely identifies oncogenic alterations for the majority of cancers, many tumors harbor no discernable driver lesion. Here, we describe the exceptional molecular phenotype of a genomically quiet kidney tumor, clear cell papillary renal cell carcinoma (CCPAP). In spite of a largely wild-type nuclear genome, CCPAP tumors exhibit severe depletion of mitochondrial DNA (mtDNA) and RNA and high levels of oxidative stress, reflecting a shift away from respiratory metabolism. Moreover, CCPAP tumors exhibit a distinct metabolic phenotype uniquely characterized by accumulation of the sugar alcohol sorbitol. Immunohistochemical staining of primary CCPAP tumor specimens recapitulates both the depletion of mtDNA-encoded proteins and a lipid-depleted metabolic phenotype, suggesting that the cytoplasmic clarity in CCPAP is primarily related to the presence of glycogen. These results argue for non-genetic profiling as a tool for the study of cancers of unknown driver.


Subject(s)
Carcinoma, Renal Cell/pathology , Cell Respiration , Kidney Neoplasms/pathology , Aerobiosis , Histocytochemistry , Humans , Immunohistochemistry , Metabolic Networks and Pathways , Oxidation-Reduction
2.
PLoS Genet ; 14(11): e1007805, 2018 11.
Article in English | MEDLINE | ID: mdl-30452458

ABSTRACT

Mitochondrial DNA (mtDNA) mutations cause severe maternally inherited syndromes and the accumulation of somatic mtDNA mutations is implicated in aging and common diseases. However, the mechanisms that influence the frequency and pathogenicity of mtDNA mutations are poorly understood. To address this matter, we created a Drosophila mtDNA mutator strain expressing a proofreading-deficient form of the mitochondrial DNA polymerase. Mutator flies have a dramatically increased somatic mtDNA mutation frequency that correlates with the dosage of the proofreading-deficient polymerase. Mutator flies also exhibit mitochondrial dysfunction, shortened lifespan, a progressive locomotor deficit, and loss of dopaminergic neurons. Surprisingly, the frequency of nonsynonymous, pathogenic, and conserved-site mutations in mutator flies exceeded predictions of a neutral mutational model, indicating the existence of a positive selection mechanism that favors deleterious mtDNA variants. We propose from these findings that deleterious mtDNA mutations are overrepresented because they selectively evade quality control surveillance or because they are amplified through compensatory mitochondrial biogenesis.


Subject(s)
DNA Polymerase gamma/genetics , DNA Polymerase gamma/metabolism , DNA, Mitochondrial/genetics , Drosophila Proteins/genetics , Drosophila Proteins/metabolism , Drosophila melanogaster/enzymology , Drosophila melanogaster/genetics , Point Mutation , Aging/genetics , Aging/metabolism , Animals , Animals, Genetically Modified , DNA Replication/genetics , Dopaminergic Neurons/metabolism , Dopaminergic Neurons/pathology , Drosophila melanogaster/cytology , Genes, Insect , Longevity/genetics , Mitochondria/enzymology , Mitochondria/genetics , Motor Activity/genetics , Organelle Biogenesis
3.
Ann Neurol ; 80(2): 301-6, 2016 08.
Article in English | MEDLINE | ID: mdl-27315116

ABSTRACT

Mitochondrial dysfunction and oxidative damage are commonly associated with early stage Alzheimer disease (AD). The accumulation of somatic mutations in mitochondrial DNA (mtDNA) has been hypothesized to be a driver of these phenotypes, but the detection of increased mutation loads has been difficult due to a lack of sensitive methods. We used an ultrasensitive next generation sequencing technique to measure the mutation load of the entire mitochondrial genome. Here, we report a significant increase in the mtDNA mutation frequency in the hippocampus of early stage AD, with the cause of these mutations being consistent with replication errors and not oxidative damage. Ann Neurol 2016;80:301-306.


Subject(s)
Alzheimer Disease/genetics , DNA, Mitochondrial/genetics , Mutation , Oxidative Stress , Aged , Aged, 80 and over , Case-Control Studies , Female , Genome, Mitochondrial/genetics , High-Throughput Nucleotide Sequencing , Hippocampus/metabolism , Humans , Male , Parietal Lobe/metabolism
4.
Brain Pathol ; 26(1): 75-81, 2016 Jan.
Article in English | MEDLINE | ID: mdl-26095919

ABSTRACT

Mortalin, an essential mitochondrial chaperone protein, has previously been implicated in the pathogenesis of a wide array of diseases, including neurodegenerative conditions such as Parkinson's disease (PD) and Alzheimer's disease. Previous reports have consistently described mortalin protein levels to be lower in the brain tissue of patients with neurodegenerative disease, with expression demonstrated to be lower in neurons of post-mortem PD brain specimens. However, to date, mortalin expression has not yet been evaluated in astrocytes of post-mortem brain tissue from either normal or PD subjects. Mortalin expression was demonstrated in mouse primary astrocyte cultures by Western blot and quantitative polymerase chain reaction (PCR). Furthermore, confocal microscopy studies in human post-mortem tissue indicated co-localization of mortalin within astrocytes. Utilizing a quantitative immunofluorescence staining approach, the protein was found to be moderately reduced (∼35%) in this cell type in the substantia nigra pars compacta, but not structures of the corpus striatum, in PD subjects as compared to age-/gender-matched controls. These findings highlight the potential contribution of disrupted astroglial function in the pathogenesis of PD.


Subject(s)
Astrocytes/metabolism , HSP70 Heat-Shock Proteins/metabolism , Mesencephalon/metabolism , Mesencephalon/pathology , Parkinson Disease/pathology , Aged , Aged, 80 and over , Animals , Animals, Newborn , Case-Control Studies , Cells, Cultured , Female , Glial Fibrillary Acidic Protein/metabolism , HSP70 Heat-Shock Proteins/genetics , Humans , Male , Mice , Middle Aged , RNA, Messenger/metabolism
5.
Neuron ; 87(2): 371-81, 2015 Jul 15.
Article in English | MEDLINE | ID: mdl-26182419

ABSTRACT

Parkinson's disease (PD) is a neurodegenerative disease caused by the loss of dopaminergic neurons in the substantia nigra. PARK2 mutations cause early-onset forms of PD. PARK2 encodes an E3 ubiquitin ligase, Parkin, that can selectively translocate to dysfunctional mitochondria to promote their removal by autophagy. However, Parkin knockout (KO) mice do not display signs of neurodegeneration. To assess Parkin function in vivo, we utilized a mouse model that accumulates dysfunctional mitochondria caused by an accelerated generation of mtDNA mutations (Mutator mice). In the absence of Parkin, dopaminergic neurons in Mutator mice degenerated causing an L-DOPA reversible motor deficit. Other neuronal populations were unaffected. Phosphorylated ubiquitin was increased in the brains of Mutator mice, indicating PINK1-Parkin activation. Parkin loss caused mitochondrial dysfunction and affected the pathogenicity but not the levels of mtDNA somatic mutations. A systemic loss of Parkin synergizes with mitochondrial dysfunction causing dopaminergic neuron death modeling PD pathogenic processes.


Subject(s)
DNA, Mitochondrial/genetics , Dopaminergic Neurons/pathology , Mitochondrial Diseases/pathology , Mutation/genetics , Substantia Nigra/pathology , Ubiquitin-Protein Ligases/metabolism , Analysis of Variance , Animals , Dopamine Plasma Membrane Transport Proteins/metabolism , Dopaminergic Neurons/drug effects , Gene Expression Regulation/drug effects , Gene Expression Regulation/genetics , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , In Vitro Techniques , Levodopa/therapeutic use , Mice , Mice, Inbred C57BL , Mice, Transgenic , Mitochondrial Diseases/genetics , Multienzyme Complexes/metabolism , Proteomics , Substantia Nigra/drug effects , Tyrosine 3-Monooxygenase/metabolism , Ubiquitin , Ubiquitin-Protein Ligases/genetics
6.
Neurobiol Aging ; 36(7): 2304-2318, 2015 Jul.
Article in English | MEDLINE | ID: mdl-25983062

ABSTRACT

Activated microglia are commonly observed in individuals with neurodegenerative disorders, including Parkinson's disease (PD) and are believed to contribute to neuronal death. This process occurs at least due partially to nicotinamide adenine dinucleotide phosphate oxidase (PHOX) activation, which leads to the production of superoxide and oxidative stress. α-Synuclein (α-Syn), a key protein implicated in PD pathogenesis, can activate microglia, contributing to death of dopaminergic neurons. Here, microglial cells (BV2) and primary cultured microglia were used to study the role that the purinergic receptor P2X7 plays in recognizing α-Syn and promoting PHOX activation. We demonstrate that both wild type and A53T mutant α-Syn readily activate PHOX, with the A53T form producing more rapid and sustained effects,that is, oxidative stress and cellular injuries. Furthermore, this process involves the activation of phosphoinositide 3-kinase (PI3K)/AKT (protein kinase B) pathway. Thus, it is concluded that stimulation of the microglial P2X7 receptor by extracellular α-Syn, with PI3K/AKT activation and increased oxidative stress, could be an important mechanism and a potential therapeutic target for PD.


Subject(s)
Enzyme Activation/genetics , Microglia/enzymology , NADPH Oxidases/metabolism , Receptors, Purinergic P2X7/physiology , alpha-Synuclein/physiology , Animals , Cell Death/genetics , Cells, Cultured , Dopaminergic Neurons/pathology , Male , Membrane Glycoproteins/metabolism , Mice, Inbred C57BL , Molecular Targeted Therapy , Mutation , NADPH Oxidase 2 , Oxidative Stress/genetics , Parkinson Disease/genetics , Parkinson Disease/therapy , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction/genetics , Signal Transduction/physiology , alpha-Synuclein/genetics
7.
Am J Pathol ; 185(2): 536-49, 2015 Feb.
Article in English | MEDLINE | ID: mdl-25482923

ABSTRACT

Mitochondrial dynamics has recently become an area of piqued interest in neurodegenerative disorders, including Parkinson disease (PD); however, the contribution of astrocytes to these disorders remains unclear. Here, we show that the level of dynamin-like protein 1 (Dlp1; official name DNM1L), which promotes mitochondrial fission, is lower in astrocytes from the brains of PD patients, and that decreased astrocytic Dlp1 likely represents a relatively early event in PD pathogenesis. In support of this conclusion, we show that Dlp1 knockdown dramatically affects mitochondrial morphological characteristics and localization in astrocytes, impairs the ability of astrocytes to adequately protect neurons from the excitotoxic effects of glutamate, and increases intracellular Ca(2+) in response to extracellular glutamate, resulting from compromised intracellular Ca(2+) buffering. Taken together, our results suggest that astrocytic mitochondrial Dlp1 is a key protein in mitochondrial dynamics and decreased Dlp1 may interfere with neuron survival in PD by disrupting Ca(2+)-coupled glutamate uptake.


Subject(s)
Calcium Signaling , Calcium/metabolism , GTP Phosphohydrolases/metabolism , Microtubule-Associated Proteins/metabolism , Mitochondria/metabolism , Mitochondrial Proteins/metabolism , Neurons/metabolism , Parkinson Disease/metabolism , Astrocytes/metabolism , Astrocytes/pathology , Cell Survival/genetics , Dynamins , Female , GTP Phosphohydrolases/genetics , Gene Knockdown Techniques , Glutamic Acid/genetics , Glutamic Acid/metabolism , Humans , Male , Microtubule-Associated Proteins/genetics , Mitochondria/genetics , Mitochondria/pathology , Mitochondrial Proteins/genetics , Neurons/pathology , Parkinson Disease/genetics , Parkinson Disease/pathology
8.
Drug Discov Today ; 18(3-4): 155-62, 2013 Feb.
Article in English | MEDLINE | ID: mdl-22982303

ABSTRACT

This review summarizes major advances in biomarker discovery for diagnosis, differential diagnosis and progression of Parkinson's disease (PD), with emphasis on neuroimaging and biochemical markers. Potential strategies to develop biomarkers capable of predicting PD in the prodromal stage before the appearance of motor symptoms or correlating with nonmotor symptoms, an active area of research, are also discussed.


Subject(s)
Biomarkers/metabolism , Parkinson Disease/diagnosis , Humans , Neuroimaging , Parkinson Disease/metabolism
9.
Alzheimers Res Ther ; 3(3): 21, 2011 Jun 29.
Article in English | MEDLINE | ID: mdl-21722346

ABSTRACT

In neurons, mitochondria serve a wide variety of processes that are integral to their function and survival. It is, therefore, not surprising that evidence of mitochondrial dysfunction is observed across numerous neurodegenerative diseases. Alzheimer's disease and Parkinson's disease are two such diseases in which aberrant mitochondrial activity is proposed to contribute to pathogenesis. Current therapies for each disease target various mechanisms, but few, if any, directly target improved mitochondrial function. Recent discoveries pertaining to mitochondrial dynamics reveal that regulation of mitochondrial fission and fusion may play a key role in the pathogenesis of these diseases and consequently could be novel future therapeutic targets.

10.
Proteomics Clin Appl ; 2(10-11): 1484-97, 2008 Oct.
Article in English | MEDLINE | ID: mdl-21136796

ABSTRACT

Proteomics has revealed itself as a powerful tool in the identification and determination of proteins and their biological significance. More recently, several groups have taken advantage of the high-throughput nature of proteomics in order to gain a more in-depth understanding of the human brain. In turn, this information has provided researchers with invaluable insight into the potential pathways and mechanisms involved in the pathogenesis of several neurodegenerative disorders, e.g., Alzheimer and Parkinson disease. Furthermore, these findings likely will improve methods to diagnose disease and monitor disease progression as well as generate novel targets for therapeutic intervention. Despite these advances, comprehensive understanding of the human brain proteome remains challenging, and requires development of improved sample enrichment, better instrumentation, and innovative analytic techniques. In this review, we will focus on the most recent progress related to identification of proteins in the human brain under normal as well as pathological conditions, mainly Alzheimer and Parkinson disease, their potential application in biomarker discovery, and discuss current advances in protein identification aimed at providing a more comprehensive understanding of the brain.

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