Chen, Pan et al. (2015) International Worm Meeting "The role of a novel genetic factor (SLC30A10) in manganese-induced toxicity in C. elegans."

Leyva-Illades, Dinorah, Aschner, Michael, Bowman, Aaron, Chen, Pan, Mukhopadhyay, Somshuvra

[International Worm Meeting, 2015]

Environmental or occupational exposure to manganese (Mn) causes a neuropathy, resembling idiopathic Parkinson's disease (PD). In cells, high Mn levels can result in oxidative stress, mitochondrial dysfunction, protein aggregation and disruption of iron homeostasis. However, the mechanisms of Mn-induced neurotoxicity remain largely unknown. Recently, newly identified mutant alleles in human SLC30A10 have been found to cause Mn accumulation in the basal ganglia. Patients with mutations in SLC30A10 show symptoms of hepatic cirrhosis, dystonia, polycythemia and hypermanganesemia. Although clinical studies indicate the potential role of SLC30A10 in cellular Mn efflux, the exact function of this protein is unknown. Here, we present novel data on the role of SLC30A10 in C. elegans. We generated transgenic animals expressing either wildtype (WT) or mutant (L89P) SLC30A10. Expression of WT SLC30A10 increased survival rate upon Mn exposure. Whereas, expression in dopaminergic (DAergic) neurons attenuated Mn-induced DAergic neurodegeneration. In both cases mutant SLC30A10 showed no protection. Moreover, WT protein also improved DAergic neuron-specific behavior upon Mn exposure, while the mutant did not. In HeLa cells and GABAergic cells, only WT protein protected against Mn-induced cell death, consistent with our results in C. elegans. The Mn level was significantly lower in HeLa cells expressing WT SLC30A10. To explore why mutant SLC30A10 loses its Mn-protecting activity, transgenic worms were generated, expressing SLC30A10 with a C-terminal green fluorescent protein (GFP) tag. We found that WT SLC30A10 localized to the cell surface, while the mutant form accumulated in the endoplasmic reticulum (ER) and cytosol. Together, these results suggest that WT SLC30A10 protects cells from Mn-induced toxicity, while the mutant protein fails to do so, likely due to its failure to localize to the cell surface. Furthermore, SLC30A10 is likely a Mn-specific transporter although it was first posited to be a zinc transporter, as it did not protect from zinc-induced lethality in C. elegans. Our results provide novel insights into the mechanisms involved in the onset of a familial form of parkinsonism and highlight the possibility of using enhanced Mn efflux as a therapeutic strategy for the management of Mn-induced parkinsonism, including that occurring as a result of mutations in SLC30A10. (supported by National Institutes of Health Grants R01-ES010563 and R00-ES020488). .

Tsur, A et al. (2011) International Worm Meeting "SUMO proteases in C. elegans."

Tsur, A, Broday, L

[International Worm Meeting, 2011]

The covalent attachment of SUMO to its target proteins is a highly dynamic and reversible post-translational modification due to the activity of SUMO proteases. Both de-conjugation of SUMO (deSUMOylation) and the processing of SUMO precursors prior to conjugation are carried out by the same family of proteases called Ubl specific proteases (ULP) in yeast, Drosophila and C. elegans and Sentrin-specific proteases (SENP) in vertebrates. ULP/SENPs are therefore key modulators of SUMOylation dynamics. In order to perform a comparative analysis of the five known C. elegans ULP proteins (ULP-1-5) we generated a phylogenic tree of over 70 different ULP/SENP sequences. The tree shows that ULP-3 which was considered to be a SUMO protease is an ortholog of the mammalian SENP8, a Nedd8-specific protease. ULP-1 appears to be the most closely related to SENP1 and SENP2. ULP-2 is highly related to the mammalian SENP6 and SENP7 suggesting a possible preference towards de-conjugation of SUMOylated species and not SUMO processing [1]. Expression of a translational reporter ULP-2::GFP is mainly nuclear and appears at late gastrulation. ulp-2(RNAi) resulted in ~30% embryonic lethality. Analysis of DLG-1 and AJM-1 fluorescence reporters showed severe defects in epidermal morphogenesis. Our findings suggest a role for ULP-2 in epithelial development. 1. Mukhopadhyay D, Dasso M. Modification in reverse: the SUMO protease. Trends Biochem Sci. 2007 Jun;32(6):286-95. Epub 2007 May 17.

Kristian Jeppsson et al. (2008) European Worm Meeting "Involvement of DAF-19 in general development and in subtype specification of cilia"

Jan Burghoorn, Filip Ystrom, Peter Swoboda, Kristian Jeppsson, Sofie Sahlin

[European Worm Meeting, 2008]

The highly conserved Caenorhabditis elegans RFX-type transcription factor. DAF-19 was originally identified as a key regulator of cilia development. (Swoboda et al. 2000). Cilia are specialized sub-cellular organelles that. in C. elegans are present exclusively at the endings of sensory neuron. dendrites. Loss of DAF-19 function leads to complete loss of all ciliated. structures in the worm. Cilia act as sensory antennae that control e.g. the. animal''s chemotactic movements in response to chemicals from the. environment. To sense many different chemical stimuli, the worm has a. variety of cilia with distinct neuron specific morphologies. Cilia with a. cylindrical morphology detect hydrophilic molecules, while adjacent wing. cilia have a fan-like structure and detect volatile odorants. How these. distinct structures and sensory modalities develop is to a large extent. unknown. For example, it is not known, which pathway determines that one. olfactory neuron develops a cylindrical ciliary structure, whereas another. forms fan-like cilia. It was hypothesized that these developmental. specifications may be accomplished by highly conserved transcription factor. cascades (Lanjuin et al. 2004; Mukhopadhyay et al. 2007).. We are interested in whether DAF-19 is not only involved in general. ciliogenesis, but also plays a role in cilia subtype specification. In. genome-wide searches for candidate direct target genes with a DAF-19. recognition site in the promoter region, the X-box, many transcription. factors were identified (Blacque et al. 2005; Efimenko et al. 2005; Chen et. al. 2006). A sizeable group of these are nuclear hormone receptors (NHR).. In C. elegans, NHR compose a large class of transcription factors (284),. some of which have been shown to function in development. We are currently. establishing expression patterns of those NHR, which are candidate direct. DAF-19 targets, including determining their DAF-19 dependence. So far we. could already demonstrate that four NHR show DAF-19 dependent expression in. ciliated sensory neurons (CSN). Interestingly, these four NHR are DAF-19. dependent in only a subset of CSN. We are continuing to analyze the. expression patterns and DAF-19 dependence of all "X-box selected" NHR and. in addition investigating their possible role in cilia subtype. specification.