Oral Sessions

Oral Sessions 細胞内・細胞間情報伝達、その他

3O4-1 Lysosomal ubiquitin ligase RNF182 regulates mTORC1 signaling through recruitment of amino acid transporter LAT1 to lysosomes Masayuki Kaneko，Xiaopeng Guo，Kazunori Imaizumi Dept Biochem, Inst Biomed & Hlth Sci, Hiroshima Univ We identified 37 ubiquitin ligases containing RING-finger and transmembrane domains. Of these, we found that RNF182, a central nervous system-specific gene, is the most highly expressed during neuronal differentiation in P19 cells and is predominantly expressed in lysosomes. To investigate its role in neural differentiation, we performed shotgun proteomics experiments and identified the lysosomal-associated transmembrane protein 4A (LAPTM4A) as its substrate. We determined that RNF182 preferentially poly-ubiquitinates LAPTM4A via K63-linked chains, suggesting that RNF182 does not promote K48 chain-mediated degradation of LAPTM4A. It was reported that LAPTM4B, from the same family as LAPTM4A, participates in recruitment of the large neutral amino acid transporter LAT1 to lysosomes, leading to leucine uptake into lysosomes and mammalian target of rapamycin complex 1 (mTORC1) activation. We showed that RNF182-mediated poly-ubiquitination of LAPTM4A facilitated the interaction of LAPTM4A and LAT1. Furthermore, we found that RNF182 expression was induced by the lysosome inhibitor chloroquine through the transcription factor TFEB. Since TFEB is inactivated by mTORC1, RNF182 may be a positive regulator of the mTORC1 pathway. Based on these results, we speculate that RNF182 upregulates mTORC1 signaling via LAPTM4A ubiquitination during neuronal differentiation.		3O4-2 Structural variations in alpha synuclein (αSyn) fibrils of Parkinson’s disease and related disorders. Kensuke Ikenaka¹，Keita Kakuda¹，Katsuya Araki¹，Cesar Aguirre²，Masatomo So²，Yuji Goto²，Hideki Mochizuki¹ ¹Dept Neurol. Grad Sch Med, Osaka Univ，²Institute of Protein Science, Osaka Univ BackgroundSynucleinopathies are neurodegenerative movement disorder characterized by the abnormal accumulation of aggregated αSyn in central nerve system. There are three main types of synucleinopathy, Parkinson’s disease (PD), diffuse Lewy bodies disease (DLBD), and multiple system atrophy (MSA). Despite of aggregations of a same protein, each three diseases has a distinct clinical phenotype as well as the different cell type specificity or region specificity in its pathology. Recent studies suggest that variations in αSyn fibril structure may related to variations in disease phenotype, in analogy to distinct prion strains that are associated with different clinical and pathological phenotypes. MethodIn this study, in order to elucidate the mechanism underlying the heterogeneities of synucleinopathies, we analyzed the characters and structures of fibrils obtained from CSF of each synucleinpathy patient. For amplification of fibrils in CSF, we employed Real-Time Quaking-Induced Conversion (RT-QuIC) technique. We amplified αSyn fibrils form 10 PD patients, 5 DLB patients, 7 MSA patients, and 10 disease control patients. Fibril structure and character were analyzed using transmission electron microscope (TEM) and the proteinase K resistance. Result and DiscussionFibril from synucleinopathy group had PK resistances compared to disease control. Especially, fibril from MSA patients had a strongest PK resistant. TEM analysis showed the distinct fibril structure in MSA patients.The data suggest that structural variations are related to the variations in synucleinopathies and amplification of fibril from CSF will be a strong tool for structure analysis of αSyn fibrils in CSF.

3O4-3 Vasopressin casts light on the suprachiasmatic nucleus Takahiro Tsuji，Chiharu Tsuji，Gareth Leng，Mike Ludwig University of Edinburgh Non-image light information, such as circadian rhythm and pupillary light reflex, is mainly received through intrinsically photosensitive retinal ganglion cells (ipRGCs) expressing melanopsin. Here we show a population of vasopressin (VP) expressing retinal ganglion cells (VP-RGCs), a part of which co-express melanopsin. Intravitreal injections of a rAAV-expressing Venus or td-to mato (under the control of the VP promoter) show that the majority of these cells project to the ventrolateral part of the suprachiasmatic nucleus (SCN). Microdialysis data show an increase in intra-SCN VP concentration in response to light stimulation of the eye more at late night than at early night. Furthermore, in response to these stimuli some SCN neurons showed an excitatory response which was blocked by icv injection of a VP V1 receptor antagonist. Light induced expression of the immediate early gene product c-Fos during the dark period was strongly reduced by prior administration of a VP V1 receptor antagonist. Thus, neurons in the ventrolateral SCN both control the output of vasopressin from the SCN by their innervation of vasopressin cells in the dorsomedial SCN, and are themselves regulated by vasopressin inputs from the retina. A well-known neurotransmitter in this pathway, glutamate, is packaged in small synaptic vesicles and can be rapidly recycled after released, while neuropeptide including VP is contained in separate, large vesicles, synthesized in cell body, and transported along the long pathway, which cannot be replenished once secreted. Accordingly, cyclical availability of neuropeptides for release may explain why a light pulse given close to the end of the night is more likely to result in a phase advance of circadian rhythms than one given earlier.