Project description:Frontotemporal lobar degeneration is a progressive neurodegenerative syndrome that is the second most common cause of early-onset dementia. Mutations in the progranulin gene are a major cause of familial frontotemporal lobar degeneration [Baker M, et al. (2006) Nature 442:916-919 and Cruts M, et al. (2006) Nature 442:920-924]. Although progranulin is involved in wound healing, inflammation, and tumor growth, its role in the nervous system and the mechanism by which insufficient levels result in neurodegeneration are poorly understood [Eriksen and Mackenzie (2008) J Neurochem 104:287-297]. We have characterized the normal function of progranulin in the nematode Caenorhabditis elegans. We found that mutants lacking pgrn-1 appear grossly normal, but exhibit fewer apoptotic cell corpses during development. This reduction in corpse number is not caused by reduced apoptosis, but instead by more rapid clearance of dying cells. Likewise, we found that macrophages cultured from progranulin KO mice displayed enhanced rates of apoptotic-cell phagocytosis. Although most neurodegenerative diseases are thought to be caused by the toxic effects of aggregated proteins, our findings suggest that susceptibility to neurodegeneration may be increased by a change in the kinetics of programmed cell death. We propose that cells that might otherwise recover from damage or injury are destroyed in progranulin mutants, which in turn facilitates disease progression.
Project description:The pathophysiology of neurodegenerative diseases is poorly understood and there are few therapeutic options. Neurodegenerative diseases are characterized by progressive neuronal dysfunction and loss, and chronic glial activation. Whether microglial activation, which is generally viewed as a secondary process, is harmful or protective in neurodegeneration remains unclear. Late-onset neurodegenerative disease observed in patients with histiocytoses, which are clonal myeloid diseases associated with somatic mutations in the RAS-MEK-ERK pathway such as BRAF(V600E), suggests a possible role of somatic mutations in myeloid cells in neurodegeneration. Yet the expression of BRAF(V600E) in the haematopoietic stem cell lineage causes leukaemic and tumoural diseases but not neurodegenerative disease. Microglia belong to a lineage of adult tissue-resident myeloid cells that develop during organogenesis from yolk-sac erythro-myeloid progenitors (EMPs) distinct from haematopoietic stem cells. We therefore hypothesized that a somatic BRAF(V600E) mutation in the EMP lineage may cause neurodegeneration. Here we show that mosaic expression of BRAF(V600E) in mouse EMPs results in clonal expansion of tissue-resident macrophages and a severe late-onset neurodegenerative disorder. This is associated with accumulation of ERK-activated amoeboid microglia in mice, and is also observed in human patients with histiocytoses. In the mouse model, neurobehavioural signs, astrogliosis, deposition of amyloid precursor protein, synaptic loss and neuronal death were driven by ERK-activated microglia and were preventable by BRAF inhibition. These results identify the fetal precursors of tissue-resident macrophages as a potential cell-of-origin for histiocytoses and demonstrate that a somatic mutation in the EMP lineage in mice can drive late-onset neurodegeneration. Moreover, these data identify activation of the MAP kinase pathway in microglia as a cause of neurodegeneration and this offers opportunities for therapeutic intervention aimed at the prevention of neuronal death in neurodegenerative diseases.
Project description:In two brothers born to consanguineous parents, we identified an unusual neurological disease that manifested with ataxia, psychomotor retardation, cerebellar and cerebral atrophy, and leukodystrophy. Via linkage analysis and exome sequencing, we identified homozygous c.2801C>T (p.(Ser934Leu)) in POLR1A (encoding RPA194, largest subunit of RNA polymerase I) and c.511C>T (p.(Arg171Trp)) in OSBPL11 (encoding oxysterol-binding protein-like protein 11). Although in silico analysis, histopathologic evidence and functional verification indicated that both variants were deleterious, segregation with the patient phenotype established that the POLR1A defect underlies the disease, as a clinically unaffected sister also was homozygous for the OSBPL11 variant. Decreased nucleolar RPA194 was observed in the skin fibroblasts of only the affected brothers, whereas intracellular cholesterol accumulation was observed in the skin biopsies of the patients and the sister homozygous for the OSBPL11 variant. Our findings provide the first report showing a complex leukodystrophy associated with POLR1A. Variants in three other RNA polymerase subunits, POLR1C, POLR3A and POLR3B, are known to cause recessive leukodystrophy similar to the disease afflicting the present family but with a later onset. Of those, POLR1C is also implicated in a mandibulofacial dysostosis syndrome without leukodystrophy as POLR1A is. This syndrome is absent in the family we present.
Project description:The pathophysiology of neurodegenerative diseases is poorly understood, and therapeutic options are few. Neurodegenerative diseases are hallmarked by progressive neuronal dysfunction and loss, associated with chronic glial and immune activation1. Microglia are the resident macrophages of the nervous system thought to be important for the clearance of debris after neuronal damage and aid to repair and regeneration. Their activation is viewed in general as a reactive process in neurodegeneration1-3. Whether microglia itself may be causative of neurodegenerative diseases is unclear. Late-onset neurodegenerative disease is frequently observed in patients with histiocytoses4-10, which are clonal myeloid diseases associated with somatic mutations in the RAS/MEK/ERK pathway such as BRAFV600E 5,11,12, suggesting a possible role of somatic mutations in neurodegeneration. Yet, expression of BRAFV600E in the hematopoietic stem cell (HSC) lineage does not cause neurodegeneration13,14. However, recent works from our laboratories and other have revealed that microglia belong to a lineage of adult tissue-resident myeloid cells that develop during organogenesis from yolk sac erythro-myeloid progenitors (EMP) distinct from HSC15-18. We thus hypothesized that somatic BRAFV600E mutations in the resident macrophage lineage may cause neurodegeneration. We found that while BRAFV600E expression in the HSC lineage causes leukemic and tumoral diseases13,14, its expression in the EMP lineage instead results in a severe late-onset neurodegenerative disorder. Neurological symptoms, neuronal death, astrogliosis, immune activation, and amyloid precursor protein deposition were driven by ERK-activated microglia clones and preventable by RAF inhibition. These results identify the fetal precursors of tissue-resident macrophages as a potential cell-of-origin for histiocytoses, and demonstrate that a somatic mutation in the EMP lineage can drive late-onset neurodegeneration. Moreover, these data identify activation of the MAP kinase pathway in microglia as a cause of neurodegenerative disease, and provide opportunities for therapeutic intervention aimed at preventing neuronal death in neurodegenerative diseases
Project description:While, transcriptional and epigenetic changes associated senescence processes are well studied, the 3D chromatin changes associated with it remains elusive. In this study, we have generated genome wide chromatin interaction maps (Hi-C), epigenetic (ChIP-Seq), replication-timing and gene expression (RNA-Seq) profiles from replication induced (RS) and oncogene induced (OIS) senescent cells. As senescence associated heterochromatin foci (SAHFs) differentiates both RS and OIS nuclei, we identified the regions that constitute SAHFs and called them Senescence Associated Heterochromatin Domains (SAHDs). Further, screening of candidate factors for SAHF induction allowed us to identify DNMT1 as a novel component that induces SAHFs by stimulation of HMGA2 expression. DNMT1 depletion does not reverse the senescence process, however, instead, depleted cells transition to a 3D genome conformation akin to that of cells in replicative senescence, suggesting that acute senescence induction (OIS) involves SAHF formation in addition to the RS-dependent 3D genome rewiring.
Project description:RNA was purified from fusiform gyrus tissue sections of autopsy-confirmed Alzheimer's cases and neurologically normal age-matched controls. The "SAM.ID" sample characteristic is a sample identifier internal to Genentech. The ID of this project in Genentech's ExpressionPlot database is PRJ0007261
Project description:Mitochondrial diseases have been shown to result from mutations in mitochondrial genes located in either the nuclear DNA (nDNA) or mitochondrial DNA (mtDNA). Mitochondrial OXPHOS complex I has 45 subunits encoded by 38 nuclear and 7 mitochondrial genes. Two male patients in a putative X-linked pedigree exhibiting a progressive neurodegenerative disorder and a severe muscle complex I enzyme defect were analyzed for mutations in the 38 nDNA and seven mtDNA encoded complex I subunits. The nDNA X-linked NDUFA1 gene (MWFE polypeptide) was discovered to harbor a novel missense mutation which changed a highly conserved glycine at position 32 to an arginine, shown to segregate with the disease. When this mutation was introduced into a NDUFA1 null hamster cell line, a substantial decrease in the complex I assembly and activity was observed. When the mtDNA of the patient was analyzed, potentially relevant missense mutations were observed in the complex I genes. Transmitochondrial cybrids containing the patient's mtDNA resulted in a mild complex I deficiency. Interestingly enough, the nDNA encoded MWFE polypeptide has been shown to interact with various mtDNA encoded complex I subunits. Therefore, we hypothesize that the novel G32R mutation in NDUFA1 is causing complex I deficiency either by itself or in synergy with additional mtDNA variants.
Project description:Uromodulin-associated kidney disease (UAKD) is caused by mutations in the uromodulin (UMOD) gene that result in a misfolded form of UMOD protein, which is normally secreted by nephrons. In UAKD patients, mutant UMOD is poorly secreted and accumulates in the ER of distal kidney epithelium, but its role in disease progression is largely unknown. Here, we modeled UMOD accumulation in mice by expressing the murine equivalent of the human UMOD p.Cys148Trp point mutation (UmodC147W/+ mice). Like affected humans, these UmodC147W/+ mice developed spontaneous and progressive kidney disease with organ failure over 24 weeks. Analysis of diseased kidneys and purified UMOD-producing cells revealed early activation of the PKR-like ER kinase/activating transcription factor 4 (PERK/ATF4) ER stress pathway, innate immune mediators, and increased apoptotic signaling, including caspase-3 activation. Unexpectedly, we also detected autophagy deficiency. Human cells expressing UMOD p.Cys147Trp recapitulated the findings in UmodC147W/+ mice, and autophagy activation with mTOR inhibitors stimulated the intracellular removal of aggregated mutant UMOD. Human cells producing mutant UMOD were susceptible to TNF-α- and TRAIL-mediated apoptosis due to increased expression of the ER stress mediator tribbles-3. Blocking TNF-α in vivo with the soluble recombinant fusion protein TNFR:Fc slowed disease progression in UmodC147W/+ mice by reducing active caspase-3, thereby preventing tubule cell death and loss of epithelial function. These findings reveal a targetable mechanism for disease processes involved in UAKD.
Project description:The human tau is a microtubule-associated intrinsically unstructured protein that forms intraneuronal cytotoxic deposits in neurodegenerative diseases, like tauopathies. Recent studies indicate that in Alzheimer's disease, ribosomal dysfunction might be a crucial event in the disease pathology. Our earlier studies had demonstrated that amorphous protein aggregation in the presence of ribosome can lead to sequestration of the ribosomal components. The present study aims at determining the effect of incubation of the full-length tau protein (Ht40) and its microtubule binding 4-repeat domain (K18) on the eukaryotic ribosome. Our in vitro studies show that incubation of Ht40 and the K18 tau variants with isolated non-translating yeast ribosome can induce a loss of ribosome physical integrity resulting in formation of tau-rRNA-ribosomal protein aggregates. Incubation with the tau protein variants also led to a disappearance of the peak indicating the ribosome profile of the HeLa cell lysate and suppression of translation in the human in vitro translation system. The incubation of tau protein with the ribosomal RNA leads to the formation of tau-rRNA aggregates. The effect of K18 on the yeast ribosome can be mitigated in the presence of cellular polyanions like heparin and tRNA, thereby indicating the electrostatic nature of the aggregation process.
Project description:Induced pluripotent stem cell (iPSC) technology can be used to model human disorders, create cell-based models of human diseases, including neurodegenerative diseases, and in establishing therapeutic strategies. To detect subtle cellular abnormalities associated with common late-onset disease in iPSCs, valid control iPSCs derived from healthy donors free of serious late-onset diseases are necessary. Here, we report the generation of iPSCs from fibroblasts obtained immediately postmortem from centenarian donors (106- and 109-years-old) who were extremely healthy until an advanced age. The iPSCs were generated using a conventional method involving OCT4, SOX2, KLF4, and c-MYC, and then differentiated into neuronal cells using a neurosphere method. The expression of molecules that play critical roles in late-onset neurodegenerative diseases by neurons differentiated from the centenarian-iPSCs was compared to that of neurons differentiated from iPSCs derived from familial Alzheimer's disease and familial Parkinson's disease (PARK4: triplication of the ? synuclein gene) patients. The results indicated that our series of iPSCs would be useful in neurodegeneration research. The iPSCs we describe, which were derived from donors with exceptional longevity who were presumed to have no serious disease risk factors, would be useful in longevity research and as valid super-controls for use in studies of various late-onset diseases.