Caenorhabditis elegans as an Experimental Model Organism to Study Parkinson's Disease-Related Genes

Parkinson’s disease (PD) is the second most common neurodegenerative disorder, mainly characterized by motor dysfunctions resulting from massive and selective loss of dopaminergic neurons. Mutations in the human parkin gene, which encodes an E3 ubiquitin ligase, are the most frequent causes of hereditary PD, leading to autosomal-recessive juvenile Parkinsonism. However, the cell biological role of Parkin and the molecular pathogenic mechanisms by which mutations cause PD are unclear. In this study, the Caenorhabditis elegans parkin ortholog, pdr-1, was identified and characterized in detail. PDR-1 is functionally conserved, since it physically associates and cooperates with enzymes of the ubiquitylation/degradation system to mediate ubiquitin conjugation. Strikingly, in contrast to pdr-1 loss-of-function mutants, the in-frame deletion mutant protein PDR-1(delaa24-247) still interacts with its co-enzymes, and moreover, the corresponding mutant pdr-1(lg103) is hypersensitive towards misfolded protein conditions. In this mutant, both cytosolic stress conferred by overexpression of mutant human alpha-synuclein, a gene linked to autosomal-dominant forms of PD, as well as endoplasmatic reticulum (ER)-derived folding stress result in severe developmental defects and lethality. Although expression of pdr-1 is regulated by all three activators of the unfolded protein response (UPR), IRE-1, PEK-1, and ATF-6, genetic analyses established a function of PDR-1 in parallel to IRE-1 signalling. Therefore, PDR-1/Parkin plays an essential role in the regulation of different proteotoxic stress pathways: it contributes to the ER-specific UPR, but also participates in the cytosolic detoxification of protein aggregates, including alpha-synuclein. The truncated protein PDR-1(delaa24-247) seems to mediate a toxic misfunction by sequestering critical components of the protein folding/degradation machinery, which is related to the stress hypersensitivity in the pdr-1(lg103) mutant. In this study, an experimental animal system was established which is well suited to identify modifiers of toxicity and relevant compounds. Such studies might allow to dissect the molecular and cellular pathways involved in the pathogenesis of PD and to identify potential therapeutic drug targets

miR-27a and miR-27b regulate autophagic clearance of damaged mitochondria by targeting PTEN-induced putative kinase 1 (PINK1)

Computational prediction of miRNA candidates for human PINK1. a Computation prediction of miRNAs expressed in human midbrain with putative binding sites in the 3′UTR of human PINK1 mRNA. We first searched miRNAs that have putative binding sites in the 3′UTR of human PINK1 mRNA by utilizing several miRNA-target prediction algorithms, such as miRanda [67], miRWalk [68], RNAhybrid [37], and Targetscan [69]. Among 49 miRNAs commonly predicted by different algorithms, 7 miRNAs were known to be expressed in human midbrain [34]. miR-27a/b are predicted to have 2 putative binding sites in the 3′UTR of human PINK1 mRNA, while all other miRNAs are predicted to have 1 putative binding site. b Computational binding prediction of miR-27a/b and their binding sites in the 3′UTR of human PINK1 mRNA. The binding free energies were determined by the RNAhybrid algorithm. (PDF 68 kb

Activation of the E3 ubiquitin ligase Parkin

Abstract The PINK1 (phosphatase and tensin homologue-induced putative kinase 1)/Parkin-dependent mitochondrial quality control pathway mediates the clearance of damaged organelles, but appears to be disrupted in Indeed, the obtained structures showed a sequential release of Parkin's intertwined domains and allowed docking of an Ub-charged E2 coenzyme, which could enable its enzymatic activity. In addition, using cellbased screening, select E2 enzymes that redundantly, cooperatively or antagonistically regulate Parkin's activation and/or enzymatic functions at different stages of the mitochondrial autophagy (mitophagy) process were identified [Fiesel et al. (2014) J. Cell Sci. 127, 3488-3504]. Other work that aims to pin-point the particular pathogenic dysfunctions of Parkin mis-sense mutations have been recently disseminated (Fabienne C. Fiesel, Thomas R. Caulfield, Elisabeth L. Moussaud-Lamodiere, Daniel F.A.R. Dourado, Kotaro Ogaki, Owen A. Ross, Samuel C. Flores, and Wolfdieter Springer, submitted). Such a structure-function approach provides the basis for the dissection of Parkin's regulation and a targeted drug design to identify small-molecule activators of this neuroprotective E3 Ub ligase

LUBAC assembles a ubiquitin signaling platform at mitochondria for signal amplification and transport of NF-κB to the nucleus

Mitochondria are increasingly recognized as cellular hubs to orchestrate signaling pathways that regulate metabolism, redox homeostasis, and cell fate decisions. Recent research revealed a role of mitochondria also in innate immune signaling; however, the mechanisms of how mitochondria affect signal transduction are poorly understood. Here, we show that the NF-κB pathway activated by TNF employs mitochondria as a platform for signal amplification and shuttling of activated NF-κB to the nucleus. TNF treatment induces the recruitment of HOIP, the catalytic component of the linear ubiquitin chain assembly complex (LUBAC), and its substrate NEMO to the outer mitochondrial membrane, where M1- and K63-linked ubiquitin chains are generated. NF-κB is locally activated and transported to the nucleus by mitochondria, leading to an increase in mitochondria-nucleus contact sites in a HOIP-dependent manner. Notably, TNF-induced stabilization of the mitochondrial kinase PINK1 furthermore contributes to signal amplification by antagonizing the M1-ubiquitin-specific deubiquitinase OTULIN. Overall, our study reveals a role for mitochondria in amplifying TNF-mediated NF-κB activation, both serving as a signaling platform, as well as a transport mode for activated NF-κB to the nuclear

LRRTM3 Interacts with APP and BACE1 and Has Variants Associating with Late-Onset Alzheimer's Disease (LOAD)

Leucine rich repeat transmembrane protein 3 (LRRTM3) is member of a synaptic protein family. LRRTM3 is a nested gene within α-T catenin (CTNNA3) and resides at the linkage peak for late-onset Alzheimer’s disease (LOAD) risk and plasma amyloid β (Aβ) levels. In-vitro knock-down of LRRTM3 was previously shown to decrease secreted Aβ, although the mechanism of this is unclear. In SH-SY5Y cells overexpressing APP and transiently transfected with LRRTM3 alone or with BACE1, we showed that LRRTM3 co-localizes with both APP and BACE1 in early endosomes, where BACE1 processing of APP occurs. Additionally, LRRTM3 co-localizes with APP in primary neuronal cultures from Tg2576 mice transduced with LRRTM3-expressing adeno-associated virus. Moreover, LRRTM3 co-immunoprecipitates with both endogenous APP and overexpressed BACE1, in HEK293T cells transfected with LRRTM3. SH-SY5Y cells with knock-down of LRRTM3 had lower BACE1 and higher CTNNA3 mRNA levels, but no change in APP. Brain mRNA levels of LRRTM3 showed significant correlations with BACE1, CTNNA3 and APP in ∼400 humans, but not in LRRTM3 knock-out mice. Finally, we assessed 69 single nucleotide polymorphisms (SNPs) within and flanking LRRTM3 in 1,567 LOADs and 2,082 controls and identified 8 SNPs within a linkage disequilibrium block encompassing 5′UTR-Intron 1 of LRRTM3 that formed multilocus genotypes (MLG) with suggestive global association with LOAD risk (p = 0.06), and significant individual MLGs. These 8 SNPs were genotyped in an independent series (1,258 LOADs and 718 controls) and had significant global and individual MLG associations in the combined dataset (p = 0.02–0.05). Collectively, these results suggest that protein interactions between LRRTM3, APP and BACE1, as well as complex associations between mRNA levels of LRRTM3, CTNNA3, APP and BACE1 in humans might influence APP metabolism and ultimately risk of AD.© 2013 Lincoln et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

Benigne perivaginale Raumforderungen: Ätiologie, Diagnostik und therapeutisches Management

Einleitung: Benigne perivaginale Raumforderungen (PVRF) sind relativ selten. Treten sie auf, stellen sie in vielen Fällen eine diagnostische und therapeutische Herausforderung dar. Vielfältige, sich oftmals überlappende Symptome, sowie ein mangelndes Bewusstsein für diese seltenen Entitäten tragen maßgeblich dazu bei. Eine inkorrekte oder verspätete Diagnose kann mit Inkontinenz, Schmerzen, Rezidiven und weiteren Komplikationen einhergehen und den Leidensweg für die betroffenen Patientinnen unnötig verlängern. In seltenen Fällen kann es zu einer malignen Transformation kommen. Ziel dieser Studie ist es, ein Bewusstsein für diese Entitäten zu schaffen sowie eine akkurate Diagnostik und Versorgung aufzuzeigen. Material und Methoden: Aus den OP-Büchern der Universitätsfrauenklinik Tübingen wurden über einen Zeitraum von fünf Jahren die Art und die Anzahl der durchgeführten urogynäkologischen Eingriffe im Allgemeinen, sowie die aufgrund einer benignen PVRF erfolgten Eingriffe im Speziellen erhoben. Aus den Krankenunterlagen wurden Diagnostik, Therapie, Histologie und postoperatives Management zusammengefasst und analysiert. Vaginale Endometriosemanifestationen fanden keine Berücksichtigung. Ergebnisse: Im Zeitraum 2011-2015 wurden an unserer Klinik insgesamt 4157 Frauen einer urogynäkologischen Operation unterzogen, 65 (1,6 %) davon aufgrund benigner PVRF. Die verschiedenen Entitäten variierten erheblich in ihrer Größe, Konfiguration und Komplexität. Die größte PVRF betrug 10 cm. PVRF traten einzeln oder multipel auf. Sie waren asymptomatisch (21,2 %) oder gingen mit einem breiten Spektrum an Symptomen einher (78,8 %). Anamnese, klinische Untersuchung, Becken-boden-Sonographie, Urethrozystoskopie und MRT waren für die Diagnostik entscheidend. In allen 65 Fällen wurde die PVRF exzidiert. In einem weiteren Fall bildete sich ein Urethradivertikel vollständig unter konservativer Therapie zurück. Fazit: Anamnese, klinische Untersuchung, Beckenboden-Sonographie, Urethrozystoskopie und MRT sind essentiell für die Diagnostik benigner PVRF. Im Falle einer Infektion sollte grundsätzlich zunächst eine konservative Therapie erfolgen. Eine komplette Exzision ist bei einem chirurgischen Vorgehen die Therapiemethode der Wahl. Das Bewusstsein für und die Vertrautheit mit den verschiedenen Entitäten ist von herausragender Bedeutung für eine korrekte Diagnose und Versorgung. Als Sekundärpathologie muss auf Divertikelsteine sowie auf eine maligne Entartung geachtet werden