Comber SD et al. (2006) Environ Pollut "Chronic toxicity of sediment-associated linear alkylbenzene sulphonates (LAS) to freshwater benthic organisms."

Hoss S, Webb S, Comber SD, Conrad AU, Marshall S

[Environ Pollut, 2006]

The toxicity of linear alkylbenzene sulphonates (LAS), to freshwater benthic organisms was assessed during exposure to spiked sediment. Lethal and sub-lethal end-points were monitored for two organisms (oligochaete Lumbriculus variegatus and nematode Caenorhabditis elegans). Results demonstrated relatively low toxicity (LOECs >100mg/kg dry weight). No observed effect concentrations (NOECs) of 81mg/kg dw (Lumbriculus) and 100mg/kg dw (Caenorhabditis) were determined. For the oligochaete, no specific endpoint was particularly sensitive to LAS. For the nematode, egg production was the most sensitive endpoint. Significant degradation was measured over the 28-day duration of the Lumbriculus study, equating to a half-life of 20days in sediment.

Mitsui Y et al. (2017) J Helminthol "Chemoattractant activity of tris-(hydroxymethyl)aminomethane for Brugia pahangi infective third-stage larvae."

Urena-Tatis KE, Aoki Y, Mitsui Y

[J Helminthol, 2017]

Urocanic acid (UCA) is known as a major chemoattractant for Strongyloides stercoralis infective third-stage larvae (L3). Since Brugia pahangi is a skin-penetrating parasitic nematode similar to S. stercoralis, UCA was expected to be a chemoattractant for B. pahangi L3. Thus, the chemoattractant activity of UCA for B. pahangi L3 was assessed. The chemotactic responses of B. pahangi L3 to UCA or acetic acid (CH3COOH) dissolved in amine solutions were assessed using an agar-plate assay. A test solution of 200 mm UCA dissolved in aqueous 270 mm tris(hydroxymethyl)aminomethane (Tris) significantly attracted B. pahangi L3 compared with deionized water (DW), while neither a solution of 200 mm UCA dissolved in aqueous 230 mm ammonia (NH3) nor 290 mm triethylamine (TEA) significantly attracted L3. Similarly, a test solution of 200 mm CH3COOH dissolved with 200 mm Tris significantly attracted L3, but neither a test solution of 200 mm CH3COOH plus 200 mm NH3 nor 200 mm TEA attracted L3. Furthermore, L3 were significantly attracted to 200 mm Tris alone, compared with DW, but avoided 200 mm NH3 and 200 mm TEA. Moreover, the chemoattractant activity of Tris for L3 was observed even at a low concentration of 25 mm, and it was observed in a mild alkaline condition but not in an acidic condition. The present study reveals that Tris is a potential chemoattractant for B. pahangi L3 while UCA is not. This finding will contribute to an understanding of the mechanisms of skin-penetrating infection of filarial L3.

Freire SL et al. (2014) J Vis Exp "Taking advantage of reduced droplet-surface interaction to optimize transport of bioanalytes in digital microfluidics."

Freire SL, Dao S, Wutkowski M, Thorne N

[J Vis Exp, 2014]

Digital microfluidics (DMF), a technique for manipulation of droplets, is a promising alternative for the development of "lab-on-a-chip" platforms. Often, droplet motion relies on the wetting of a surface, directly associated with the application of an electric field; surface interactions, however, make motion dependent on droplet contents, limiting the breadth of applications of the technique. Some alternatives have been presented to minimize this dependence. However, they rely on the addition of extra chemical species to the droplet or its surroundings, which could potentially interact with droplet moieties. Addressing this challenge, our group recently developed Field-DW devices to allow the transport of cells and proteins in DMF, without extra additives. Here, the protocol for device fabrication and operation is provided, including the electronic interface for motion control. We also continue the studies with the devices, showing that multicellular, relatively large, model organisms can also be transported, arguably unaffected by the electric fields required for device operation.

Dong B et al. (2007) J Biol Chem "Ca(2+)/calmodulin directly interacts with the pleckstrin homology domain of AKT1."

Dong B, Valencia CA, Liu R

[J Biol Chem, 2007]

AKT kinase, also known as protein kinase B, is a key regulator of cell growth, proliferation and metabolism. The activation of the AKT signaling pathway is one of the most frequent molecular alterations in a wide variety of human cancers. Dickson and coworkers recently observed that Ca2+/calmodulin (Ca2+/CaM) may be a common regulator of AKT1 activation (1). In our efforts to scan the mRNA-displayed proteome libraries for Ca2+/CaM-binding proteins, we found that both human and C. elegans AKT1 kinases bound to CaM in a Ca2+-dependent manner (2,3). Here we demonstrated that Ca2+/CaM and human AKT1 were efficiently co-immunoprecipitated and their interaction was direct rather than mediated by other proteins. The binding is in part attributed to the first 42 residues of the PH domain, a region that is critical for the recognition of its lipid ligands. The PH domain of human AKT1 can disrupt the complex of the full-length AKT1 with Ca2+/CaM. In addition, Ca2+/CaM competes with PtdIns(3,4,5)P3 for interaction with the PH domain of human AKT1. Our findings suggest that Ca2+/CaM is directly involved in regulating the functions of AKT1, presumably by releasing the activated AKT1 from the plasma membrane and/or prohibiting it from re-association with phosphoinostides on plasma membrane.

Mukherjee S et al. (2017) Alzheimers Dement "Systems biology approach to late-onset Alzheimer's disease genome-wide association study identifies novel candidate genes validated using brain expression data and Caenorhabditis elegans experiments."

Burgess JD, Carr DT, Dickson DW, Naj AC, Crane PK, Kauwe JS, Russell JC, Serie D, Mukherjee S, Ertekin-Taner N, Younkin MA, Boehme KL, Kaeberlein MR, Fardo DW, Montine TJ

[Alzheimers Dement, 2017]

INTRODUCTION: We sought to determine whether a systems biology approach may identify novel late-onset Alzheimer's disease (LOAD) loci. METHODS: We performed gene-wide association analyses and integrated results with human protein-protein interaction data using network analyses. We performed functional validation on novel genes using a transgenic Caenorhabditis elegans A proteotoxicity model and evaluated novel genes using brain expression data from people with LOAD and other neurodegenerative conditions. RESULTS: We identified 13 novel candidate LOAD genes outside chromosome 19. Of those, RNA interference knockdowns of the C. elegans orthologs of UBC, NDUFS3, EGR1, and ATP5H were associated with A toxicity, and NDUFS3, SLC25A11, ATP5H, and APP were differentially expressed in the temporal cortex. DISCUSSION: Network analyses identified novel LOAD candidate genes. We demonstrated a functional role for four of these in a C. elegans model and found enrichment of differentially expressed genes in the temporal cortex.

Dickey CA et al. (2006) J Neurosci "Deletion of the ubiquitin ligase CHIP leads to the accumulation, but not the aggregation, of both endogenous phospho- and caspase-3-cleaved tau species."

Dickey CA, Patterson C, Caldwell KA, Eckman C, Petrucelli L, Lee WC, Cao S, Dunmore JH, Zehr C, Lin WL, Hutton M, West G, Dickson DW, Caldwell GA, Yue M, Clark AM

[J Neurosci, 2006]

Accumulation of the microtubule-associated protein tau into neurofibrillary lesions is a pathological consequence of several neurodegenerative diseases, including Parkinson''s disease and Alzheimer''s disease. Hereditary mutations in the MAPT gene were shown to promote the formation of structurally distinct tau aggregates in patients that had a parkinsonian-like clinical presentation. Whether tau aggregates themselves or the soluble intermediate species that precede their aggregation are neurotoxic entities in these disorders has yet to be resolved; however, recent in vivo evidence supports the latter. We hypothesized that depletion of CHIP, a tau ubiquitin ligase, would lead to an increase in abnormal tau. Here, we show that deletion of CHIP in mice leads to the accumulation of non-aggregated, ubiquitin-negative, hyperphosphorylated tau species. CHIP-/- mice also have increased neuronal caspase-3 levels and activity, as well as caspase-cleaved tau immunoreactivity. Overexpression of mutant (P301L) human tau in CHIP-/- mice is insufficient to promote either argyrophilic or "pre-tangle" structures, despite marked phospho-tau accumulation throughout the brain. These observations are supported in post-developmental studies using RNA interference for CHIP (chn-1) in Caenorhabditis elegans and cell culture systems. Our results demonstrate that CHIP is a primary component in the ubiquitin-dependent degradation of tau. We also show that hyperphosphorylation and caspase-3 cleavage of tau both occur before aggregate formation. Based on these findings, we propose that polyubiquitination of tau by CHIP may facilitate the formation of insoluble filamentous tau lesions.

DEVKOTA, RANJAN et al. (2015) International Worm Meeting "Novel regulators of the C. elegans UNC-6/Netrin pathway."

DEVKOTA, RANJAN, Garriga, Gian, Morck, Catarina, CHIEN, JASON

[International Worm Meeting, 2015]

Precise targeting of axons is essential for the proper functioning of the nervous system. Axonal growth cones respond to various guidance cues and navigate through a complex environment to reach their synaptic targets1. Even though many guidance cues and receptors have been identified, there is less knowledge of how axons respond to different cues in different phases of their growth. The aim of this project is to enhance the understanding of axon guidance by focusing on the development of the bilaterally symmetric Hermaphrodite Specific Neurons (HSNs). Each HSN extends a single axon to the ventral nerve cord, which then turns anteriorly and extends to the nerve ring2. The HSNs innervate vulval muscles and the AVF neurons to stimulate hermaphrodite egg laying and locomotion, respectively3,4. We have isolated three mutants whose HSN axons fail to extend ventrally. Our preliminary mapping, complementation and genetic interaction data suggest that these mutations define three genes that function together to antagonize the conserved UNC-6/Netrin signaling pathway. Our initial goal is to identify the underlying mutations with whole genome sequencing and rescue experiments. The identification of these genes will shed light on how the UNC-6/Netrin pathway is modulated for proper axon guidance along the dorsal-ventral axis.References:1. Dickson, B. J. Molecular mechanisms of axon guidance. Science 298, 1959-1964(2002).2. Garriga, G., Desai, C. & Horvitz, H. R. Cell interactions control the direction of outgrowth, branching and fasciculation of the HSN axons of Caenorhabditis elegans. Development 117, 1071-1087(1993).3. Desai, C., Garriga, G., McIntire, S. L. & Horvitz, H. R. A genetic pathway for the development of the Caenorhabditis elegans HSN motor neurons. Nature 336, 638-646(1988).4. Hardaker, L. A., Singer, E., Kerr, R., Zhou, G. & Schafer, W. R. Serotonin modulates locomotory behavior and coordinates egg-laying and movement in Caenorhabditis elegans. J. Neurobiol. 49, 303-313(2001). .

Murakami T et al. (2015) Neuron "ALS/FTD Mutation-Induced Phase Transition of FUS Liquid Droplets and Reversible Hydrogels into Irreversible Hydrogels Impairs RNP Granule Function."

Vendruscolo M, Favrin G, Kronenberg-Versteeg D, Holt C, Murakami T, Dodd RB, Kaminski CF, Costa AR, Schierle GS, Meadows W, St George-Hyslop P, Dong L, Michel CH, Rees E, Wakutani Y, Fraser PE, Yang SP, Li Y, Lin JQ, Qamar S, Ron D, Tartaglia GG, Miyashita A, Chan FT, Shneider NA, Ferry RR, Lin WL, Dickson DW, Zhen M, Schmitt-Ulms G

[Neuron, 2015]

The mechanisms by which mutations in FUS and other RNA binding proteins cause ALS and FTD remain controversial. We propose a model in which low-complexity (LC) domains of FUS drive its physiologically reversible assembly into membrane-free, liquid droplet and hydrogel-like structures. ALS/FTD mutations in LC or non-LC domains induce further phase transition into poorly soluble fibrillar hydrogels distinct from conventional amyloids. These assemblies are necessary and sufficient for neurotoxicity in a C. elegans model of FUS-dependent neurodegeneration. They trap other ribonucleoprotein (RNP) granule components and disrupt RNP granule function. One consequence is impairment of new protein synthesis by cytoplasmic RNP granules in axon terminals, where RNP granules regulate local RNA metabolism and translation. Nuclear FUS granules may be similarly affected. Inhibiting formation of these fibrillar hydrogel assemblies mitigates neurotoxicity and suggests a potential therapeutic strategy that may also be applicable to ALS/FTD associated with mutations in other RNA binding proteins.

Currey, Heather N et al. (2020) MicroPubl Biol

Liachko, Nicole F, Currey, Heather N, Malinkevich, Anna, Melquist, Penny

[MicroPubl Biol, 2020]

Aggregates of the protein tau are the hallmark of several neurodegenerative diseases including Alzheimers disease, frontotemporal lobar degeneration (FTLD-tau), progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), Picks disease, and chronic traumatic encephalopathy (CTE) (VandeVrede, Boxer et al. 2020). Mutations in the gene coding for tau, MAPT, can cause FTLD-tau, directly linking tau dysfunction with disease (Dickson, Kouri et al. 2011). Another protein, TDP-43, comprises aggregates which are the primary hallmark of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD-TDP), and mutations in the gene coding for TDP-43, TARDBP, can cause disease (Kawakami, Arai et al. 2019). To model tau or TDP-43 proteinopathies, transgenic C. elegans have been generated that express the full-length human protein pan-neuronally. These worms exhibit significant uncoordinated movement on plates and impaired thrashing in liquid (Kraemer, Zhang et al. 2003; Liachko, Guthrie et al. 2010). However, tau- and TDP-43- expressing worms are not paralyzed; they still move their heads and have some motility on the plate (coiling, crawling with tail-drag, head swinging) which are not captured in standard crawling or thrashing assays. To assay differences in total activity, we used a WMicroTracker ARENA System (Phylumtech, AR and InVivo Biosystems, USA). The ARENA captures population level activity data by relying on optical interferometry, which uses a large array of infrared LED microbeams to detect both the movement and position of worms on a culture plate. Disruption of an LED microbeam by worm movement is recorded by repeat scans of the 6-well culture plate, and allows for real-time processing. The software identifies changes in the location of disrupted beams between scans and assigns an activity score based on differences identified between each consecutive scan (Simonetta SH). Both tau- and TDP-43- expressing worms had significantly less activity per minute than N2 (Figure 1). Further, we found the ARENA- assessed activity data recapitulated the relative severity of phenotypes among the strains as measured by motility assays. For example, both CK10 (tau V337M) and CK144 (tau WT) have significantly uncoordinated movement when crawling or thrashing in liquid, with CK144 having worse motility than CK10, due to its much higher burden of total tau protein expressed (Kraemer, Zhang et al. 2003). Likewise, CK410 (TDP-43 WT) worms have slightly impaired motility compared with N2 when crawling on a plate, CK423 (TDP-43 M337V) are severely uncoordinated, and CK426 (TDP-43 A315T) have the most severe uncoordinated phenotype. The relative toxicities of these strains stem from the effects of the mutations, as TDP-43 protein expression is relatively even among these transgenic strains (Liachko, Guthrie et al. 2010). Interestingly, the ARENA captures activity of these severely uncoordinated worms that move poorly in motility assays such as crawling on an NGM plate or thrashing in liquid (Kraemer, Zhang et al. 2003; Liachko, Guthrie et al. 2010). Therefore, ARENA assessment of aggregate activity may be a more accurate metric for capturing non-locomotor movement of C. elegans that are severely uncoordinated.

Kramer NJ et al. (2016) Science "Spt4 selectively regulates the expression of C9orf72 sense and antisense mutant transcripts."

Van Blitterswijk M, Petersen RC, Parisi JE, Prudencio M, Couthouis J, Dickson DW, Jansen-West K, Josephs KA, Petrucelli L, Bonini NM, Gitler AD, Tabassian LJ, Rademakers R, Carlomagno Y, Feng Y, Boylan K, Gao FB, Kramer NJ, Belzil V, Daughrity L, Link CD, Graff-Radford NR, Cohen SN, Knopman DS, Deng N, Chew J, Boeve BF, Garrett A, Pregent L, Paul JW, Goodman LD, Gendron TF, Cook CN, Almeida S, Cheng TH, Zhang YJ

[Science, 2016]

An expanded hexanucleotide repeat in C9orf72 causes amyotrophic lateral sclerosis and frontotemporal dementia (c9FTD/ALS). Therapeutics are being developed to target RNAs containing the expanded repeat sequence (GGGGCC); however, this approach is complicated by the presence of antisense strand transcription of expanded GGCCCC repeats. We found that targeting the transcription elongation factor Spt4 selectively decreased production of both sense and antisense expanded transcripts, as well as their translated dipeptide repeat (DPR) products, and also mitigated degeneration in animal models. Knockdown of SUPT4H1, the human Spt4 ortholog, similarly decreased production of sense and antisense RNA foci, as well as DPR proteins, in patient cells. Therapeutic targeting of a single factor to eliminate c9FTD/ALS pathological features offers advantages over approaches that require targeting sense and antisense repeats separately.