Brian Kraemer et al. (2007) International Worm Meeting "SUT-1 Enables tau neurotoxicity."

Brian Kraemer, Gerard Schellenberg

[International Worm Meeting, 2007]

A number of neurodegenerative disorders have insoluble deposits of pathological tau including Alzheimer''s disease (AD), Down syndrome, corticobasal degeneration, progressive supranuclear palsy, Picks disease, Guam amyotrophic lateral sclerosis/Parkinsons dementia complex and frontotemporal dementia with parkinsonism chromosome 17 type (FTDP-17). Mutations in tau cause FTDP-17 demonstrating that tau abnormalities can cause neurodegeneration. We have generated transgenic C. elegans that express human tau to model tau induced neurodegeneration. Using a classical genetic approach we have isolated genes called suppressor of tau (sut) genes that when mutated alleviate the neurotoxic effects of tau. The recessive mutation, sut-1(bk79) suppresses the Unc phenotype, tau aggregation, and neurodegenerative changes caused by tau. Cloning of sut-1 revealed that it encodes a short protein with no known protein domains. sut-1 mutants lack detectable SUT-1 protein. Using yeast two hybrid and in vitro binding assays, we demonstrated that SUT-1 interacts with the worm Enabled homolog, a regulator of cytoskeletal dynamics. However, the mechanism of sut-1 mediated suppression of tau neurotoxicity remains unclear. SUT-1 protein shares little sequence homology with known human proteins. The molecular function of SUT-1 is unknown. Since genetic ablation of SUT-1 prevents tauopathy in our worm model, pharmacological inhibition of any existing human analogs of SUT-1 may be viable targets for intervention in human tauopathy disorders.

Brian C Kraemer et al. (2003) International Worm Meeting "Modeling tauopathy in Caenorhabditis elegans"

Brian C Kraemer, Gerard D Schellenberg, James H Thomas

[International Worm Meeting, 2003]

Aggregated tau protein accumulates in the brain lesions of human neurodegernative diseases such as frontotemporal dementia with parkinsonism chromosome 17 type (FTDP-17) and Alzheimers disease. FTDP-17 is caused by mutations in MAPT, the gene encoding tau. We transgenically expressed normal and FTDP-17 mutant human tau in Caenorhabditis elegans to model tauopathy disorders. Tau transgenes caused a progressive behavioral defect linked to subsequent accumulations of insoluble phosphorylated tau and neurodegeneration. Animals expressing FTDP-17 mutant tau were more impaired than those expressing normal tau. Cholinergic neurons showed defective pre-synaptic but not post-synaptic function. This worm model for human disease is presently being used to explore the genetic requirements of tau-induced neurodegeneration and formation of pathological tau through genetic screening. An F1 screen of approximately 13,000,000 ethylnitrosurea (ENU) mutagenized haploid genomes for dominant suppressors of the tau-induced Unc phenotype yielded 54 putative dominant suppressor alleles. Of these 54, 19 bred true. Secondary screening to remove suppressors with reduced transgene expression or transgene silencing revealed that 6 of 19 lines expressed levels of tau similar to the parental non-mutagenized line. Additionally, an F2 screen was carried out. Approximately 50,000 haploid genomes were screened for recessive mutations yielding 72 putative F2 suppressors. Of these 72, 11 bred true. Secondary screening for tau expression resulted in 3 of 11 lines that expressed levels of tau similar to the parental non-mutagenized line. Efforts to map and clone these mutations are underway.

Brian C Kraemer et al. (2005) International Worm Meeting "Using C. elegans to identify genes that prevent pathological effects of human tau."

Brian C Kraemer, James H Thomas, Gerard D Schellenberg

[International Worm Meeting, 2005]

Alzheimers disease (AD) is the most common neurodegenerative disease. The brains of AD patients exhibit two primary pathological hallmarks, neuritic plaques and the neurofibrillary tangles. The microtubule binding protein tau is the primary constituent of neurofibrillary tangles. Mutations in the human gene encoding tau cause the neurodegenerative disorder, frontotemporal dementia with parkinsonism chromosome 17 type (FTDP-17), demonstrating that tau defects can cause disease. Neurofibrillary tangles composed of insoluble hyperphosphorylated tau protein are characteristic of Alzheimers disease, FTDP-17, and many other diseases with pathological tau. Such disorders with pathological tau protein are collectively referred to as tauopathies. We previously reported a C. elegans model for tauopathy using normal and FTDP-17 mutant tau cDNAs as transgenes expressed in worm neurons. To investigate possible mechanisms of tau-induced neurodegeneration, we conducted a genome wide screen in tau transgenic C. elegans. We identified a number of genes that enhance the tau induced phenotype using the feeding RNA mediated interference (RNAi) procedure and the RNAi library of Kamath et al1. Genome wide RNAi screening revealed 75 genes that enhance the tau induced phenotype. These modifier genes fall into a number of broad classes including members of kinase, chaperone, protease, and phosphatase gene families. Forty-six of these worm tauopathy modifier genes share homology to known human genes while 29 are specific to nematodes. We tested a variety of Unc mutants on tau RNAi enhancer clones; 11 clones modified a variety of different Unc phenotypes while 64 were specific to the tau induced Unc phenotype. To confirm the RNAi findings, we introduced the tau transgene into the background of genetic loss of function mutations for a subset of the identified tau enhancer genes. Tau transgenic animals homozygous for loss of acr-14, aex-1, or lin-44 each exhibited impaired motility relative to the tau transgene alone. Furthermore these mutations caused increased accumulation of insoluble tau consistent with the exacerbation of the uncoordinated phenotype. This work suggests a number of novel potential candidate genes involved in preventing tau toxicity, as well as identifying genes previously implicated in tauopathy. 1. Kamath et al. Nature 421(6920), 231-7. 2003.

Chen X et al. (2015) Mol Neurodegener "Erratum to: Ethosuximide ameliorates neurodegenerative disease phenotypes by modulating DAF-16/FOXO target gene expression."

McCue HV, Chen X, Morgan A, Kashyap SS, Barclay JW, Kraemer BC, Burgoyne RD, Wong SQ

[Mol Neurodegener, 2015]

The original version of this article [1] unfortunately contained a mistake. The author list contained a spelling error for the author Hannah V. McCue. The original article has been corrected for this error. The corrected author list is given below:Xi Chen, Hannah V. McCue, Shi Quan Wong, Sudhanva S. Kashyap, Brian C. Kraemer, Jeff W. Barclay, Robert D. Burgoyne and Alan Morgan

Liachko, Nicole et al. (2015) International Worm Meeting "Using stress assays to define aberrant neuronal signaling in a C. elegans model of TDP-43 proteinopathy."

Kraemer, Brian, Liachko, Nicole, Kushleika, John, Bird, Thomas

[International Worm Meeting, 2015]

Accumulation of the protein TDP-43 in neuronal aggregates is the major pathological feature of two devastating neurodegenerative diseases, amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTLD-TDP). TDP-43 is an essential protein and is involved in many aspects of RNA metabolism from transcription to translation, although its most critical contribution may be regulation of the majority of cellular mRNA splicing events. Disruption of the normal activity of TDP-43, either by mutation of TDP-43 or alterations in TDP-43 regulatory pathways, promotes neuronal dysfuction and degeneration in a variety of model systems including C. elegans, Drosophila, zebrafish, mammalian cell culture, and mice. To study the cellular, molecular, and genetic underpinnings of TDP-43 mediated neurotoxicity in a tractable model system, we have developed C. elegans models of TDP-43 proteinopathy expressing either wild type or disease-causing mutant TDP-43 pan-neuronally (TDP-43 tg). These transgenic animals display early, progressive motor dysfunction, decreased lifespan, and age-dependent degeneration of specific types of neurons, including GABA-ergic and dopaminergic neurons (1). However, not all TDP-43 expressing neurons undergo apparent neurodegeneration, indicating differences in sensitivities of specific populations of neurons to the presence of TDP-43. Surveying the range of responses to TDP-43 may provide a set of shared characteristics for populations of neurons susceptible or resistant to aberrant TDP-43. To investigate neuronal function in TDP-43 tg animals, we are utilizing a panel of behavioral, stress response, and stress survival assays. Results from these assays have identified individual sensory neurons with functional impairments in the absence of early neuronal cell body degeneration. These TDP-43 sensitive neurons will allow dissection of the effects of neurotoxic TDP-43 on neuronal function, and provide insight into the upstream processes leading to TDP-43 dependent neurodegeneration. 1. N. F. Liachko, C. R. Guthrie, B. C. Kraemer, Phosphorylation Promotes Neurotoxicity in a Caenorhabditis elegans Model of TDP-43 Proteinopathy. J Neurosci 30, 16208-16219 (2010).

Kow, Rebecca et al. (2015) International Worm Meeting "Identification of Dopamine Signaling Pathway Components that Modify Tau Neurotoxicity in Tau Transgenic Caenorhabditis elegans."

Kraemer, Brian, Wheeler, Jeanna, Kow, Rebecca

[International Worm Meeting, 2015]

The human protein tau can accumulate into toxic aggregates seen in multiple neurodegenerative diseases such as Alzheimer's disease and frontotemporal lobar degeneration. Expression of human tau in Caenorhabditis elegans causes significantly reduced locomotor ability, tau aggregation, progressive loss of neurons, and shortened lifespan. We have previously shown that loss of dop-2 and dop-3 dopamine receptors significantly ameliorated tau transgene-induced phenotypes and reduced accumulation of toxic forms of tau. To determine the key signaling components that link dopamine receptor activity to tau regulation, we generated C. elegans strains containing the tau transgene and loss of function mutations in known dopamine signaling pathway genes. Candidate modifier genes span the entire pathway, from dopamine metabolism enzymes to the ultimate downstream transcriptional regulators. We are currently assaying these worms for enhancement or suppression of tau transgene-induced locomotor dysfunction. One candidate enhancer of tau transgene-induced locomotor dysfunction identified so far is egl-8. Identified enhancers and suppressors of tau transgene-induced locomotor phenotypes will be examined for neuron loss, lifespan changes, and toxic tau accumulation.

Saxton, Aleen et al. (2013) International Worm Meeting "A C. elegans genome-wide RNAi screen identifies modifiers of mutant TDP-43."

Kraemer, Brian, Saxton, Aleen, Liachko, Nicole

[International Worm Meeting, 2013]

Aggregated, hyperphosphorylated TAR DNA-binding protein-43 (TDP-43) occurs in multiple neurodegenerative diseases including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration. Transgenic C. elegans expressing familial ALS mutant TDP-43 (ckIs423[Psnb-1::hTDP-43(M337V)] subsequently, TDP-43M337V) have a variety of abnormal phenotypes including uncoordinated locomotion. Using the Ahringer RNA interference (RNAi) feeding library, we have completed a genome-wide screen for suppressors or enhancers of TDP-43M337V motor defects and fitness in an eri-1(mg366);lin-15(n744) RNAi enhancing mutant background. Out of the 16,750 unique RNAi clones in the library, a total of 2,486 potential positive candidates were selected for retesting following the initial screen. Positive candidates have also been tested in non-transgenic C. elegans to confirm TDP-43 specific effects of the RNAi on movement and viability. From these candidates, we have identified many candidate suppressors that improved TDP-43M337V movement, and many candidate enhancers that worsened TDP-43M337V phenotypes, causing paralysis, developmental arrest, or death. Validation of hits from the collection of identified RNAi modifiers is ongoing using genetic loss of function studies in candidate genes quantitatively in the TDP-43M337V transgenic animals. Our screening approach promises to uncover conserved genes involved in TDP-43 related neurotoxicity.

Guthrie, Chris R. et al. (2009) International Worm Meeting "SUT-2 potentiates tau-induced neurotoxicity in C. elegans."

Kraemer, Brian C., Guthrie, Chris R.

[International Worm Meeting, 2009]

We have used our transgenic C. elegans model of human tauopathy diseases by expressing human tau in worm neurons to explore pathways that contribute to tau-induced neurodegeneration. Expression of mutated human tau causes uncoordinated locomotion (Unc) in C. elegans. This animal recapitulates produces several hallmarks of human tauopathies including accumulation of detergent insoluble phosphorylated tau protein and neurodegeneration. We carried out a forward genetic screen for mutations that prevent the tau-induced Unc phenotype in order to identify genes that control tau neurotoxicity. Recessive loss of function mutations in the sut-2 locus suppresses the Unc phenotype, tau aggregation, and neurodegenerative changes caused by human tau. We cloned the sut-2 gene and found it encodes a novel sub-type of CCCH zinc finger protein conserved across animal phyla. SUT-2 shares significant identity with the mammalian SUT-2 (MSUT-2). To identify SUT-2 interacting proteins, we conducted a yeast two hybrid screen and found SUT-2 binds to ZYG-12, the sole C. elegans HOOK protein family member. Likewise, SUT-2 binds ZYG-12 in in vitro protein binding assays. Furthermore, loss of ZYG-12 leads to a marked upregulation of SUT-2 protein supporting the connection between SUT-2 and ZYG-12. Of the three human orthologs of ZYG-12, MSUT-2 binds only to HOOK2. This suggests the interaction between SUT-2 and HOOK family proteins is conserved across animal phyla. Immunofluorescent microscopy of endogenous MSUT-2 revealed that MSUT-2 localizes nuclear domains known as nuclear speckles or interchromatin granule clusters. These nuclear sub domains are known to contain components of the pre-mRNA splicing machinery. The identification of sut-2 as a gene required for tau neurotoxicity in C. elegans may suggest new neuroprotective strategies capable of arresting tau pathogenesis in human tauopathy disease.

Liachko, Nicole et al. (2009) International Worm Meeting "A C. elegans model of ALS and other TDP-43 proteinopathy disorders."

Kraemer, Brian, Guthrie, Chris, Liachko, Nicole

[International Worm Meeting, 2009]

Aging related neurodegenerative disorders are frequently characterized by lesions containing deposits of insoluble protein aggregates. Recently, the lesions seen in the degenerating neurons of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with ubiquitin positive inclusions (FTLD-U) were discovered to consist primarily of the TDP-43 protein. Likewise TDP-43 inclusions are also seen in many cases of other diseases leading to dementia including Alzheimer''s disease (AD), Parkinson''s disease (PD), dementia with Lewy bodies (DLB), and Guam amyotrophic lateral sclerosis/Parkinson''s dementia complex (ALS/PDC). Disorders with deposits of TDP-43 are now referred to as TDP-43 proteinopathy disorders. Of these, ALS is of particular interest because there is a causal link between TDP-43 and ALS. TDP-43 mutations cause some forms of familial ALS proving that abnormal TDP-43 can be a cause of neurodegeneration, although the mechanism by which TDP-43 abnormalities lead to neurodegeneration is poorly understood. To model TDP-43 neurotoxicity, we have transgenically expressed human TDP-43 protein in the neurons of C. elegans. Neuronal expression of normal human TDP-43 causes a relatively mild Unc phenotype, while expression of ALS causing mutant forms of TDP-43 leads to a robust Unc phenotype. Consistent with the mild Unc phenotype, we see little if any neurodegeneration caused by the wild type TDP-43 transgene. However, ALS mutant TDP-43 expressing transgenic animals exhibit dramatic degeneration of motor neurons. As in authentic human TDP-43 proteinopathy disorders, we observe the accumulation of insoluble TDP-43 protein in our model, both for wild type and mutant TDP-43 transgenes. This is in keeping with the observation that wild type TDP-43 forms inclusions in most sporadic ALS cases, while mutant TDP-43 is known to form aggregates in familial ALS cases. Detailed characterization of TDP-43 associated phenotypes and analysis of the genes and pathways contributing to TDP-43 mediated neurodegeneration are underway. The long term goal of this work is to develop neuroprotective strategies for neurodegenerative disorders with TPD-43 protein deposits.

Liachko, Nicole F. et al. (2009) International Worm Meeting "Functional characterization of the C. elegans homologs of human neurodegenerative disease proteins TDP-43 and PGRN."

Kraemer, Brian C., Liachko, Nicole F.

[International Worm Meeting, 2009]

Neurodegenerative diseases affect a growing percentage of the human population as average lifespan increases. It is of great importance to characterize the causative agents involved in these diseases, as these will provide clues for treatments or disease-delaying interventions. Recent studies have identified the TAR DNA binding protein-43 (TDP-43) to be the pathological protein involved in a large percentage of cases of frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). Loss of function mutations in the progranulin (PGRN) gene have been linked to TDP-43-pathological FTLD, as PGRN mutants accumulate TDP-43 neuronal aggregates. In addition, cell culture studies have shown that PGRN may mediate caspase-dependent cleavage of TDP-43, but it remains to be determined the exact nature of the interaction between PGRN and TDP-43 as it relates to FTLD. It is essential to characterize the normal and pathological functions of PGRN and TDP-43 to identify possible interventions in FTLD and ALS. To this end, we are testing the C. elegans TDP-43 and PGRN homologs tdp-1 and pgrn-1. These genes have 39% and 55% homology, respectively, to their human counterparts, and are the primary homologous genes identified following protein-protein BLAST using either the C. elegans or human sequences as queries. Since the protein sequences are highly conserved between the human and C. elegans TDP-43/tdp-1 and PGRN/pgrn-1, it is possible that their functions are conserved as well, and observations made using C. elegans may complement research in mammalian systems. We are characterizing the behavior and physiology of mutant tdp-1 and pgrn-1 to provide clues into the normal functions of these genes, including observations of movement, responses to stimuli, and neuronal development. In addition, we are performing microarrays to determine which gene expression levels are affected by tdp-1 or pgrn-1 mutations, and are comparing differentially regulated genes between the two mutants for further insight into their normal functions and functional interactions. In the long term, this work will provide a C. elegans system to study evolutionarily conserved functions of the human disease genes TDP-43 and PGRN, providing tools for understanding the mechanisms of FTLD.