start-ver=1.4 cd-journal=joma no-vol=10 cd-vols= no-issue=1 article-no= start-page=4798 end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2020 dt-pub=20200316 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Development of an experimental method of systematically estimating protein expression limits in HEK293 cells en-subtitle= kn-subtitle= en-abstract= kn-abstract=Protein overexpression sometimes causes cellular defects, although the underlying mechanism is still unknown. A protein's expression limit, which triggers cellular defects, is a useful indication of the underlying mechanism. In this study, we developed an experimental method of estimating the expression limits of target proteins in the human embryonic kidney cell line HEK293 by measuring the proteins' expression levels in cells that survived after the high-copy introduction of plasmid DNA by which the proteins were expressed under a strong cytomegalovirus promoter. The expression limits of nonfluorescent target proteins were indirectly estimated by measuring the levels of green fluorescent protein (GFP) connected to the target proteins with the self-cleaving sequence P2A. The expression limit of a model GFP was similar to 5.0% of the total protein, and sustained GFP overexpression caused cell death. The expression limits of GFPs with mitochondria-targeting signals and endoplasmic reticulum localization signals were 1.6% and 0.38%, respectively. The expression limits of four proteins involved in vesicular trafficking were far lower compared to a red fluorescent protein. The protein expression limit estimation method developed will be valuable for defining toxic proteins and consequences of protein overexpression. en-copyright= kn-copyright= en-aut-name=MoriYoshihiro en-aut-sei=Mori en-aut-mei=Yoshihiro kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=YoshidaYuki en-aut-sei=Yoshida en-aut-mei=Yuki kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=MoriyaHisao en-aut-sei=Moriya en-aut-mei=Hisao kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= affil-num=1 en-affil=Graduate School of Natural Science and Technology, Okayama University kn-affil= affil-num=2 en-affil=Sony Computer Science Laboratories kn-affil= affil-num=3 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=4 en-affil=Research Core for Interdisciplinary Sciences, Okayama University kn-affil= en-keyword=Biological techniques kn-keyword=Biological techniques en-keyword=Cell biology kn-keyword=Cell biology en-keyword=Gene expression analysis kn-keyword=Gene expression analysis en-keyword=Molecular biology kn-keyword=Molecular biology en-keyword=Protein translocation kn-keyword=Protein translocation en-keyword=Protein transport kn-keyword=Protein transport END start-ver=1.4 cd-journal=joma no-vol=12 cd-vols= no-issue=5 article-no= start-page= end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2020 dt-pub=20200516 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Knockout of MMP3 Weakens Solid Tumor Organoids and Cancer Extracellular Vesicles en-subtitle= kn-subtitle= en-abstract= kn-abstract=The tumor organoid (tumoroid) model in three-dimensional (3D) culture systems has been developed to reflect more closely the in vivo tumors than 2D-cultured tumor cells. Notably, extracellular vesicles (EVs) are efficiently collectible from the culture supernatant of gel-free tumoroids. Matrix metalloproteinase (MMP) 3 is a multi-functional factor playing crucial roles in tumor progression. However, roles of MMP3 within tumor growth and EVs have not unveiled. Here, we investigated the protumorigenic roles of MMP3 on integrities of tumoroids and EVs. We generated MMP3-knockout (KO) cells using the CRISPR/Cas9 system from rapidly metastatic LuM1 tumor cells. Moreover, we established fluorescent cell lines with palmitoylation signal-fused fluorescent proteins (tdTomato and enhanced GFP). Then we confirmed the exchange of EVs between cellular populations and tumoroids. LuM1-tumoroids released large EVs (200-1000 nm) and small EVs (50-200 nm) while the knockout of MMP3 resulted in the additional release of broken EVs from tumoroids. The loss of MMP3 led to a significant reduction in tumoroid size and the development of the necrotic area within tumoroids. MMP3 and CD9 (a category-1 EV marker tetraspanin protein) were significantly down-regulated in MMP3-KO cells and their EV fraction. Moreover, CD63, another member of the tetraspanin family, was significantly reduced only in the EVs fractions of the MMP3-KO cells compared to their counterpart. These weakened phenotypes of MMP3-KO were markedly rescued by the addition of MMP3-rich EVs or conditioned medium (CM) collected from LuM1-tumoroids, which caused a dramatic rise in the expression of MMP3, CD9, and Ki-67 (a marker of proliferating cells) in the MMP3-null/CD9-low tumoroids. Notably, MMP3 enriched in tumoroids-derived EVs and CM deeply penetrated recipient MMP3-KO tumoroids, resulting in a remarkable enlargement of solid tumoroids, while MMP3-null EVs did not. These data demonstrate that EVs can mediate molecular transfer of MMP3, resulting in increasing the proliferation and tumorigenesis, indicating crucial roles of MMP3 in tumor progression. en-copyright= kn-copyright= en-aut-name=TahaEman A. en-aut-sei=Taha en-aut-mei=Eman A. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=SogawaChiharu en-aut-sei=Sogawa en-aut-mei=Chiharu kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=OkushaYuka en-aut-sei=Okusha en-aut-mei=Yuka kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=KawaiHotaka en-aut-sei=Kawai en-aut-mei=Hotaka kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=OoMay Wathone en-aut-sei=Oo en-aut-mei=May Wathone kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=ElseoudiAbdellatif en-aut-sei=Elseoudi en-aut-mei=Abdellatif kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=LuYanyin en-aut-sei=Lu en-aut-mei=Yanyin kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=NagatsukaHitoshi en-aut-sei=Nagatsuka en-aut-mei=Hitoshi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= en-aut-name=KubotaSatoshi en-aut-sei=Kubota en-aut-mei=Satoshi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=9 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=10 ORCID= en-aut-name=OkamotoKuniaki en-aut-sei=Okamoto en-aut-mei=Kuniaki kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=11 ORCID= en-aut-name=EguchiTakanori en-aut-sei=Eguchi en-aut-mei=Takanori kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=12 ORCID= affil-num=1 en-affil=Department of Dental Pharmacology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=2 en-affil=Department of Dental Pharmacology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=3 en-affil=Department of Dental Pharmacology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=4 en-affil=Department of Oral Pathology and Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=5 en-affil=Department of Oral Pathology and Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=6 en-affil=Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=7 en-affil=Department of Dental Pharmacology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=8 en-affil=Department of Oral Pathology and Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=9 en-affil=Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=10 en-affil=Department of Medical Bioengineering, Okayama University Graduate School of Natural Science and Technology kn-affil= affil-num=11 en-affil=Department of Dental Pharmacology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=12 en-affil=Department of Dental Pharmacology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= en-keyword=matrix metalloproteinase 3 (MMP3) kn-keyword=matrix metalloproteinase 3 (MMP3) en-keyword=extracellular vesicles (EVs) kn-keyword=extracellular vesicles (EVs) en-keyword=tumoroid kn-keyword=tumoroid en-keyword=tumor organoid kn-keyword=tumor organoid en-keyword=tumorigenesis kn-keyword=tumorigenesis en-keyword=three-dimensional (3D) culture system kn-keyword=three-dimensional (3D) culture system END start-ver=1.4 cd-journal=joma no-vol=12 cd-vols= no-issue=1 article-no= start-page=82 end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2019 dt-pub=20191229 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Hematopoietic Cells Derived from Cancer Stem Cells Generated from Mouse Induced Pluripotent Stem Cells en-subtitle= kn-subtitle= en-abstract= kn-abstract=Cancer stem cells (CSCs) represent the subpopulation of cancer cells with the ability to differentiate into other cell phenotypes and initiated tumorigenesis. Previously, we reported generating CSCs from mouse induced pluripotent stem cells (miPSCs). Here, we investigated the ability of the CSCs to differentiate into hematopoietic cells. First, the primary cells were isolated from malignant tumors that were formed by the CSCs. Non-adherent cells (NACs) that arose from adherent cells were collected and their viability, as well as the morphology and expression of hematopoietic cell markers, were analyzed. Moreover, NACs were injected into the tail vein of busulfan conditioned Balb/c nude mice. Finally, CSCs were induced to differentiate to macrophages while using IL3 and SCF. The round nucleated NACs were found to be viable, positive for hematopoietic lineage markers and CD34, and expressed hematopoietic markers, just like homing to the bone marrow. When NACs were injected into mice, Wright-Giemsa staining showed that the number of white blood cells got higher than those in the control mice after four weeks. CSCs also showed the ability to differentiate toward macrophages. CSCs were demonstrated to have the potential to provide progenies with hematopoietic markers, morphology, and homing ability to the bone marrow, which could give new insight into the tumor microenvironment according to the plasticity of CSCs. en-copyright= kn-copyright= en-aut-name=HassanGhmkin en-aut-sei=Hassan en-aut-mei=Ghmkin kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=AfifySaid M. en-aut-sei=Afify en-aut-mei=Said M. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=NairNeha en-aut-sei=Nair en-aut-mei=Neha kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=KumonKazuki en-aut-sei=Kumon en-aut-mei=Kazuki kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=OsmanAmira en-aut-sei=Osman en-aut-mei=Amira kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=DuJuan en-aut-sei=Du en-aut-mei=Juan kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=MansourHager en-aut-sei=Mansour en-aut-mei=Hager kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=Abu QuoraHagar A. en-aut-sei=Abu Quora en-aut-mei=Hagar A. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= en-aut-name=NawaraHend M. en-aut-sei=Nawara en-aut-mei=Hend M. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=9 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=10 ORCID= en-aut-name=ZahraMaram H. en-aut-sei=Zahra en-aut-mei=Maram H. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=11 ORCID= en-aut-name=OkadaNobuhiro en-aut-sei=Okada en-aut-mei=Nobuhiro kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=12 ORCID= en-aut-name=SenoAkimasa en-aut-sei=Seno en-aut-mei=Akimasa kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=13 ORCID= en-aut-name=SenoMasaharu en-aut-sei=Seno en-aut-mei=Masaharu kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=14 ORCID= affil-num=1 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=2 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=3 en-affil=Graduate School of Natural Science and Technology, Okayama University kn-affil= affil-num=4 en-affil=Graduate School of Natural Science and Technology, Okayama University kn-affil= affil-num=5 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=6 en-affil=Graduate School of Natural Science and Technology, Okayama University kn-affil= affil-num=7 en-affil= kn-affil=Graduate School of Natural Science and Technology, Okayama University affil-num=8 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=9 en-affil=Graduate School of Natural Science and Technology, Okayama University kn-affil= affil-num=10 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=11 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=12 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=13 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=14 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= en-keyword=Induced pluripotent stem cells kn-keyword=Induced pluripotent stem cells en-keyword=Cancer stem cells differentiation kn-keyword=Cancer stem cells differentiation en-keyword=tumor microenvironment kn-keyword=tumor microenvironment en-keyword=hematopoietic cells kn-keyword=hematopoietic cells END start-ver=1.4 cd-journal=joma no-vol=9 cd-vols= no-issue= article-no= start-page=8866 end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2019 dt-pub=2019620 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Yeast screening system reveals the inhibitory mechanism of cancer cell proliferation by benzyl isothiocyanate through down-regulation of Mis12 en-subtitle= kn-subtitle= en-abstract= kn-abstract=Benzyl isothiocyanate (BITC) is a naturally-occurring isothiocyanate derived from cruciferous vegetables. BITC has been reported to inhibit the proliferation of various cancer cells, which is believed to be important for the inhibition of tumorigenesis. However, the detailed mechanisms of action remain unclear. In this study, we employed a budding yeast Saccharomyces cerevisiae as a model organism for screening. Twelve genes including MTW1 were identified as the overexpression suppressors for the antiproliferative effect of BITC using the genome-wide multi-copy plasmid collection for S. cerevisiae. Overexpression of the kinetochore protein Mtw1 counteracts the antiproliferative effect of BITC in yeast. The inhibitory effect of BITC on the proliferation of human colon cancer HCT-116 cells was consistently suppressed by the overexpression of Mis12, a human orthologue of Mtw1, and enhanced by the knockdown of Mis12. We also found that BITC increased the phosphorylated and ubiquitinated Mis12 level with consequent reduction of Mis12, suggesting that BITC degrades Mis12 through an ubiquitin-proteasome system. Furthermore, cell cycle analysis showed that the change in the Mis12 level affected the cell cycle distribution and the sensitivity to the BITC-induced apoptosis. These results provide evidence that BITC suppresses cell proliferation through the post-transcriptional regulation of the kinetochore protein Mis12. en-copyright= kn-copyright= en-aut-name=Abe-KanohNaomi en-aut-sei=Abe-Kanoh en-aut-mei=Naomi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=KunisueNarumi en-aut-sei=Kunisue en-aut-mei=Narumi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=MyojinTakumi en-aut-sei=Myojin en-aut-mei=Takumi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=ChinoAyako en-aut-sei=Chino en-aut-mei=Ayako kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=MunemasaShintaro en-aut-sei=Munemasa en-aut-mei=Shintaro kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=MurataYoshiyuki en-aut-sei=Murata en-aut-mei=Yoshiyuki kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=MoriyaHisao en-aut-sei=Moriya en-aut-mei=Hisao kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= en-aut-name=NakamuraYoshimasa en-aut-sei=Nakamura en-aut-mei=Yoshimasa kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=9 ORCID= affil-num=1 en-affil=Graduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=2 en-affil=Graduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=3 en-affil=Graduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=4 en-affil=Research Core for Interdisciplinary Sciences, Okayama University kn-affil= affil-num=5 en-affil=Graduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=6 en-affil=Graduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=7 en-affil= Graduate School of Natural Science and Technology, Okayama University kn-affil= affil-num=8 en-affil=Research Core for Interdisciplinary Sciences, Okayama University kn-affil= affil-num=9 en-affil=Graduate School of Environmental and Life Science, Okayama University, Okayama kn-affil= END start-ver=1.4 cd-journal=joma no-vol=7 cd-vols= no-issue= article-no= start-page=160 end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2019 dt-pub=20190827 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=The Golgin Protein Giantin Regulates Interconnections Between Golgi Stacks en-subtitle= kn-subtitle= en-abstract= kn-abstract= Golgins are a family of Golgi-localized long coiled-coil proteins. The major golgin function is thought to be the tethering of vesicles, membranes, and cytoskeletal elements to the Golgi. We previously showed that knockdown of one of the longest golgins, Giantin, altered the glycosylation patterns of cell surfaces and the kinetics of cargo transport, suggesting that Giantin maintains correct glycosylation through slowing down transport within the Golgi. Giantin knockdown also altered the sizes and numbers of mini Golgi stacks generated by microtubule de-polymerization, suggesting that it maintains the independence of individual Golgi stacks. Therefore, it is presumed that Golgi stacks lose their independence following Giantin knockdown, allowing easier and possibly increased transport among stacks and abnormal glycosylation. To gain structural insights into the independence of Golgi stacks, we herein performed electron tomography and 3D modeling of Golgi stacks in Giantin knockdown cells. Compared with control cells, Giantin-knockdown cells had fewer and smaller fenestrae within each cisterna. This was supported by data showing that the diffusion rate of Golgi membrane proteins is faster in Giantin-knockdown Golgi, indicating that Giantin knockdown structurally and functionally increases connectivity among Golgi cisternae and stacks. This increased connectivity suggests that contrary to the cis-golgin tether model, Giantin instead inhibits the tether and fusion of nearby Golgi cisternae and stacks, resulting in transport difficulties between stacks that may enable the correct glycosylation of proteins and lipids passing through the Golgi. en-copyright= kn-copyright= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=Hayashi-NishinoMitsuko en-aut-sei=Hayashi-Nishino en-aut-mei=Mitsuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=ShakunoTakuto en-aut-sei=Shakuno en-aut-mei=Takuto kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=MasudaJunko en-aut-sei=Masuda en-aut-mei=Junko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=KoreishiMayuko en-aut-sei=Koreishi en-aut-mei=Mayuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=MurakamiRuna en-aut-sei=Murakami en-aut-mei=Runa kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=NakamuraYoshimasa en-aut-sei=Nakamura en-aut-mei=Yoshimasa kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=NakamuraToshiyuki en-aut-sei=Nakamura en-aut-mei=Toshiyuki kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= en-aut-name=Abe-KanohNaomi en-aut-sei=Abe-Kanoh en-aut-mei=Naomi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=9 ORCID= en-aut-name=HonjoYasuko en-aut-sei=Honjo en-aut-mei=Yasuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=10 ORCID= en-aut-name=MalsamJoerg en-aut-sei=Malsam en-aut-mei=Joerg kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=11 ORCID= en-aut-name=YuSidney en-aut-sei=Yu en-aut-mei=Sidney kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=12 ORCID= en-aut-name=ishinoKunihiko en-aut-sei=ishino en-aut-mei=Kunihiko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=13 ORCID= affil-num=1 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=2 en-affil=Institute of Scientific and Industrial Research, Osaka University kn-affil= affil-num=3 en-affil=Graduate School of Natural Science and Technology, Okayama University kn-affil= affil-num=4 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=5 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=6 en-affil=Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University kn-affil= affil-num=7 en-affil=raduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=8 en-affil=raduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=9 en-affil=raduate School of Environmental and Life Science, Okayama University kn-affil= affil-num=10 en-affil=esearch Institute for Radiation Biology and Medicine, Hiroshima University kn-affil= affil-num=11 en-affil=Center for Biochemistry (BZH), Heidelberg University kn-affil= affil-num=12 en-affil=School of Biomedical Sciences, The Chinese University of Hong Kong kn-affil= affil-num=13 en-affil=Institute of Scientific and Industrial Research, Osaka University kn-affil= en-keyword=Golgi kn-keyword=Golgi en-keyword=golgins kn-keyword=golgins en-keyword=glycosylation kn-keyword=glycosylation en-keyword=endoplasmic reticulum kn-keyword=endoplasmic reticulum en-keyword=electron tomography kn-keyword=electron tomography END start-ver=1.4 cd-journal=joma no-vol=25 cd-vols= no-issue=146 article-no= start-page=241 end-page=244 dt-received= dt-revised= dt-accepted= dt-pub-year=2013 dt-pub=201311 dt-online= en-article= kn-article= en-subject= kn-subject= en-title=ゴルジ体内の輸送:糖タンパク質の移動に関する研究 kn-title=Transport within the Golgi: for the Study of Glycoprotein Movement en-subtitle= kn-subtitle= en-abstract= kn-abstract= en-copyright= kn-copyright= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name=佐藤あやの kn-aut-sei=佐藤 kn-aut-mei=あやの aut-affil-num=1 ORCID= en-aut-name=IwakiriYasuko en-aut-sei=Iwakiri en-aut-mei=Yasuko kn-aut-name=岩切泰子 kn-aut-sei=岩切 kn-aut-mei=泰子 aut-affil-num=2 ORCID= affil-num=1 en-affil= kn-affil=岡山大学大学院自然科学研究科 affil-num=2 en-affil= kn-affil=エール大学医学部内科消化器科 en-keyword=Golgi kn-keyword=Golgi en-keyword=glycosylation kn-keyword=glycosylation en-keyword=intracellular trafficking kn-keyword=intracellular trafficking END start-ver=1.4 cd-journal=joma no-vol=182 cd-vols= no-issue=3 article-no= start-page=786 end-page=795 dt-received= dt-revised= dt-accepted= dt-pub-year=2013 dt-pub=201303 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Absence of Nogo-B (Reticulon 4B) Facilitates Hepatic Stellate Cell Apoptosis and Diminishes Hepatic Fibrosis in Mice en-subtitle= kn-subtitle= en-abstract= kn-abstract=Nogo-B (reticulon 4B) accentuates hepatic fibrosis and cirrhosis, but the mechanism remains unclear. The aim of this study was to identify the role of Nogo-B in hepatic stellate cell (HSC) apoptosis in cirrhotic livers. Cirrhosis was generated by carbon tetrachloride inhalation in wild-type (WT) and Nogo-A/B knockout (Nogo-B KO) mice. HSCs were isolated from WT and Nogo-B KO mice and cultured for activation and transformation to myofibroblasts (MF-HSCs). Human hepatic stellate cells (LX2 cells) were used to assess apoptotic responses of activated HSCs after silencing or overexpressing Nogo-B. Livers from cirrhotic Nogo-B KO mice showed significantly reduced fibrosis (P < 0.05) compared with WT mice. Apoptotic cells were more prominent in fibrotic areas of cirrhotic Nogo-B KO livers. Nogo-B KO MF-HSCs showed significantly increased Levels of apoptotic markers, cleaved poly (ADP-ribose) polymerase, and caspase-3 and -8 (P < 0.05) compared with WT MF-HSCs in response to staurosporine. Treatment with tunicamycin, an endoplasmic reticulum stress inducer, increased cleaved caspase-3 and -8 levels in Nogo-B KO MF-HSCs compared with WT MF-HSCs (P < 0.01). In LX2 cells, Nogo-B knockdown enhanced apoptosis in response to staurosporine, whereas Nogo-B overexpression inhibited apoptosis. The absence of Nogo-B enhances apoptosis of HSCs in experimental cirrhosis. Selective blockade of Nogo-B in HSCs may represent a potential therapeutic strategy to mitigate liver fibrosis. (Am J Pathol 2013, 182: 786-795; http://dx.doLorg/10.1016Aajpath.2012.11.032) en-copyright= kn-copyright= en-aut-name=TashiroKeitaro en-aut-sei=Tashiro en-aut-mei=Keitaro kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=UtsumiTeruo en-aut-sei=Utsumi en-aut-mei=Teruo kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=ChungChuhan en-aut-sei=Chung en-aut-mei=Chuhan kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=IwakiriYasuko en-aut-sei=Iwakiri en-aut-mei=Yasuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= affil-num=1 en-affil= kn-affil=Yale Univ, Sch Med, Sect Digest Dis, Dept Internal Med affil-num=2 en-affil= kn-affil=Okayama Univ, Grad Sch Nat Sci affil-num=3 en-affil= kn-affil=Yale Univ, Sch Med, Sect Digest Dis, Dept Internal Med affil-num=4 en-affil= kn-affil=Yale Univ, Sch Med, Sect Digest Dis, Dept Internal Med affil-num=5 en-affil= kn-affil=Yale Univ, Sch Med, Sect Digest Dis, Dept Internal Med END start-ver=1.4 cd-journal=joma no-vol=8 cd-vols= no-issue=3 article-no= start-page= end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2013 dt-pub=20130521 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=The Golgin Tether Giantin Regulates the Secretory Pathway by Controlling Stack Organization within Golgi Apparatus en-subtitle= kn-subtitle= en-abstract= kn-abstract=Golgins are coiled-coil proteins that play a key role in the regulation of Golgi architecture and function. Giantin, the largest golgin in mammals, forms a complex with p115, rab1, GM130, and soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), thereby facilitating vesicle tethering and fusion processes around the Golgi apparatus. Treatment with the microtubule destabilizing drug nocodazole transforms the Golgi ribbon into individual Golgi stacks. Here we show that siRNA-mediated depletion of giantin resulted in more dispersed Golgi stacks after nocodazole treatment than by control treatment, without changing the average cisternal length. Furthermore, depletion of giantin caused an increase in cargo transport that was associated with altered cell surface protein glycosylation. Drosophila S2 cells are known to have dispersed Golgi stacks and no giantin homolog. The exogenous expression of mammalian giantin cDNA in S2 cells resulted in clustered Golgi stacks, similar to the Golgi ribbon in mammalian cells. These results suggest that the spatial organization of the Golgi ribbon is mediated by giantin, which also plays a role in cargo transport and sugar modifications. en-copyright= kn-copyright= en-aut-name=KoreishiMayuko en-aut-sei=Koreishi en-aut-mei=Mayuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=GniadekThomas J. en-aut-sei=Gniadek en-aut-mei=Thomas J. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=YuSidney en-aut-sei=Yu en-aut-mei=Sidney kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=MasudaJunko en-aut-sei=Masuda en-aut-mei=Junko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=HonjoYasuko en-aut-sei=Honjo en-aut-mei=Yasuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= affil-num=1 en-affil= kn-affil=Okayama Univ, Grad Sch Nat Sci & Technol affil-num=2 en-affil= kn-affil=Johns Hopkins Univ, Sch Med, Dept Pathol affil-num=3 en-affil= kn-affil=Chinese Univ Hong Kong, Sch Biomed Sci, Shatin affil-num=4 en-affil= kn-affil=NIAID, Mucosal Immun Sect, Lab Host Def affil-num=5 en-affil= kn-affil=Okayama Univ, RCIS affil-num=6 en-affil= kn-affil=Okayama Univ, Grad Sch Nat Sci & Technol END start-ver=1.4 cd-journal=joma no-vol=8 cd-vols= no-issue=1 article-no= start-page= end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2013 dt-pub=20130118 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=CK2 Phosphorylates Sec31 and Regulates ER-To-Golgi Trafficking en-subtitle= kn-subtitle= en-abstract= kn-abstract=Protein export from the endoplasmic reticulum (ER) is an initial and rate-limiting step of molecular trafficking and secretion. This is mediated by coat protein II (COPII)-coated vesicles, whose formation requires small GTPase Sar1 and 6 Sec proteins including Sec23 and Sec31. Sec31 is a component of the outer layer of COPII coat and has been identified as a phosphoprotein. The initiation and promotion of COPII vesicle formation is regulated by Sar1; however, the mechanism regulating the completion of COPII vesicle formation followed by vesicle release is largely unknown. Hypothesizing that the Sec31 phosphorylation may be such a mechanism, we identified phosphorylation sites in the middle linker region of Sec31. Sec31 phosphorylation appeared to decrease its association with ER membranes and Sec23. Non-phosphorylatable mutant of Sec31 stayed longer at ER exit sites and bound more strongly to Sec23. We also found that CK2 is one of the kinases responsible for Sec31 phosphorylation because CK2 knockdown decreased Sec31 phosphorylation, whereas CK2 overexpression increased Sec31 phosphorylation. Furthermore, CK2 knockdown increased affinity of Sec31 for Sec23 and inhibited ER-to-Golgi trafficking. These results suggest that Sec31 phosphorylation by CK2 controls the duration of COPII vesicle formation, which regulates ER-to-Golgi trafficking. en-copyright= kn-copyright= en-aut-name=KoreishiMayuko en-aut-sei=Koreishi en-aut-mei=Mayuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=YuSidney en-aut-sei=Yu en-aut-mei=Sidney kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=OdaMayumi en-aut-sei=Oda en-aut-mei=Mayumi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=HonjoYasuko en-aut-sei=Honjo en-aut-mei=Yasuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= affil-num=1 en-affil= kn-affil=Okayama Univ, Grad Sch Nat Sci & Technol affil-num=2 en-affil= kn-affil=Chinese Univ Hong Kong, Sch Biomed Sci affil-num=3 en-affil= kn-affil=Okayama Univ, Grad Sch Nat Sci & Technol affil-num=4 en-affil= kn-affil=Okayama Univ, Res Core Interdisciplinary Sci RCIS affil-num=5 en-affil= kn-affil=Okayama Univ, Grad Sch Nat Sci & Technol END start-ver=1.4 cd-journal=joma no-vol=7 cd-vols= no-issue=4 article-no= start-page= end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2012 dt-pub=20120412 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=A Model of Cancer Stem Cells Derived from Mouse Induced Pluripotent Stem Cells en-subtitle= kn-subtitle= en-abstract= kn-abstract=Cancer stem cells (CSCs) are capable of continuous proliferation and self-renewal and are proposed to play significant roles in oncogenesis, tumor growth, metastasis and cancer recurrence. CSCs are considered derived from normal stem cells affected by the tumor microenvironment although the mechanism of development is not clear yet. In 2007, Yamanaka's group succeeded in generating Nanog mouse induced pluripotent stem (miPS) cells, in which green fluorescent protein (GFP) has been inserted into the 5'-untranslated region of the Nanog gene. Usually, iPS cells, just like embryonic stem cells, are considered to be induced into progenitor cells, which differentiate into various normal phenotypes depending on the normal niche. We hypothesized that CSCs could be derived from Nanog miPS cells in the conditioned culture medium of cancer cell lines, which is a mimic of carcinoma microenvironment. As a result, the Nanog miPS cells treated with the conditioned medium of mouse Lewis lung carcinoma acquired characteristics of CSCs, in that they formed spheroids expressing GFP in suspension culture, and had a high tumorigenicity in Balb/c nude mice exhibiting angiogenesis in vivo. In addition, these iPS-derived CSCs had a capacity of self-renewal and expressed the marker genes, Nanog, Rex1, Eras, Esg1 and Cripto, associated with stem cell properties and an undifferentiated state. Thus we concluded that a model of CSCs was originally developed from miPS cells and proposed the conditioned culture medium of cancer cell lines might perform as niche for producing CSCs. The model of CSCs and the procedure of their establishment will help study the genetic alterations and the secreted factors in the tumor microenvironment which convert miPS cells to CSCs. Furthermore, the identification of potentially bona fide markers of CSCs, which will help the development of novel anti-cancer therapies, might be possible though the CSC model. en-copyright= kn-copyright= en-aut-name=ChenLing en-aut-sei=Chen en-aut-mei=Ling kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=KasaiTomonari en-aut-sei=Kasai en-aut-mei=Tomonari kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=LiYueguang en-aut-sei=Li en-aut-mei=Yueguang kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=SugiiYuh en-aut-sei=Sugii en-aut-mei=Yuh kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=JinGuoliang en-aut-sei=Jin en-aut-mei=Guoliang kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=OkadaMasashi en-aut-sei=Okada en-aut-mei=Masashi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=VaidyanathArun en-aut-sei=Vaidyanath en-aut-mei=Arun kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=MizutaniAkifumi en-aut-sei=Mizutani en-aut-mei=Akifumi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=9 ORCID= en-aut-name=KudohTakayuki en-aut-sei=Kudoh en-aut-mei=Takayuki kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=10 ORCID= en-aut-name=HendrixMary J. C. en-aut-sei=Hendrix en-aut-mei=Mary J. C. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=11 ORCID= en-aut-name=SalomonDavid S en-aut-sei=Salomon en-aut-mei=David S kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=12 ORCID= en-aut-name=FuLi en-aut-sei=Fu en-aut-mei=Li kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=13 ORCID= en-aut-name=SenoMasaharu en-aut-sei=Seno en-aut-mei=Masaharu kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=14 ORCID= affil-num=1 en-affil= kn-affil=Okayama Univ affil-num=2 en-affil= kn-affil=Okayama Univ affil-num=3 en-affil= kn-affil=Tianjin 4th Ctr Hosp affil-num=4 en-affil= kn-affil=Okayama Univ affil-num=5 en-affil= kn-affil=Okayama Univ affil-num=6 en-affil= kn-affil=Okayama Univ affil-num=7 en-affil= kn-affil=Okayama Univ affil-num=8 en-affil= kn-affil=Okayama Univ affil-num=9 en-affil= kn-affil=Okayama Univ affil-num=10 en-affil= kn-affil=Okayama Univ affil-num=11 en-affil= kn-affil=Northwestern Univ affil-num=12 en-affil= kn-affil=NCI affil-num=13 en-affil= kn-affil=Tianjin Med Univ affil-num=14 en-affil= kn-affil=Okayama Univ END start-ver=1.4 cd-journal=joma no-vol=2011 cd-vols= no-issue=1 article-no= start-page=5 end-page=11 dt-received= dt-revised= dt-accepted= dt-pub-year=201- dt-pub=2010809 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Detection of in situ cleaved p115 with the cut specific antibodies in rapid protein inactivation system by tobacco etch viral protease cleavage en-subtitle= kn-subtitle= en-abstract= kn-abstract=Gene perturbation methods are commonly used in the study of gene and protein function. The authors of this paper recently developed a rapid protein inactivation technique utilizing tobacco etch virus (TEV)-derived protease. TEV protease recognizes the ENLYFQG (Glu-Asn-Leu-Tyr-Phe-Gln-Gly) amino acid sequence and specifically cleaves between Q and G. The authors developed antibodies that recognize the cleaved TEV (ENLYFQ) sequence, both in vitro and in vivo, but do not bind to uncleaved TEV (ENLYFQG). Using these antibodies, in situ protein cleavage was successfully detected. These antibodies used in combination with the TEV protease may be a useful complement to other perturbation methods. en-copyright= kn-copyright= en-aut-name=KoreishiMayuko en-aut-sei=Koreishi en-aut-mei=Mayuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=HonjoYasuko en-aut-sei=Honjo en-aut-mei=Yasuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= affil-num=1 en-affil= kn-affil=The Research Core for Interdisciplinary Sciences (RCIS), Okayama University affil-num=2 en-affil= kn-affil=The Research Core for Interdisciplinary Sciences (RCIS), Okayama University affil-num=3 en-affil= kn-affil=The Research Core for Interdisciplinary Sciences (RCIS), Okayama University en-keyword=TEV protease kn-keyword=TEV protease en-keyword=Golgi kn-keyword=Golgi en-keyword=golgins kn-keyword=golgins en-keyword=microinjection kn-keyword=microinjection en-keyword=recombinant proteins kn-keyword=recombinant proteins END start-ver=1.4 cd-journal=joma no-vol=7 cd-vols= no-issue=1 article-no= start-page= end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2012 dt-pub=20120118 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=TRAPPC9 Mediates the Interaction between p150Glued and COPII Vesicles at the Target Membrane en-subtitle= kn-subtitle= en-abstract= kn-abstract=Background: The transport of endoplasmic reticulum (ER)-derived COPII vesicles toward the ER-Golgi intermediate compartment (ERGIC) requires cytoplasmic dynein and is dependent on microtubules. p150Glued, a subunit of dynactin, has been implicated in the transport of COPII vesicles via its interaction with COPII coat components Sec23 and Sec24. However, whether and how COPII vesicle tether, TRAPP (Transport protein particle), plays a role in the interaction between COPII vesicles and microtubules is currently unknown. Principle Findings: We address the functional relationship between COPII tether TRAPP and dynactin. Overexpressed TRAPP subunits interfered with microtubule architecture by competing p150Glued away from the MTOC. TRAPP subunit TRAPPC9 bound directly to p150Glued via the same carboxyl terminal domain of p150Glued that binds Sec23 and Sec24. TRAPPC9 also inhibited the interaction between p150Glued and Sec23/Sec24 both in vitro and in vivo, suggesting that TRAPPC9 serves to uncouple p150Glued from the COPII coat, and to relay the vesicle-dynactin interaction at the target membrane. Conclusions: These findings provide a new perspective on the function of TRAPP as an adaptor between the ERGIC membrane and dynactin. By preserving the connection between dynactin and the tethered and/or fused vesicles, TRAPP allows nascent ERGIC to continue the movement along the microtubules as they mature into the cis-Golgi. en-copyright= kn-copyright= en-aut-name=ZongMin en-aut-sei=Zong en-aut-mei=Min kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=YuMei Kuen en-aut-sei=Yu en-aut-mei=Mei Kuen kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=SiuKa Yu en-aut-sei=Siu en-aut-mei=Ka Yu kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=NgWing Yan en-aut-sei=Ng en-aut-mei=Wing Yan kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=ChanHsiao Chang en-aut-sei=Chan en-aut-mei=Hsiao Chang kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=TannerJulian A. en-aut-sei=Tanner en-aut-mei=Julian A. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=YuSidney en-aut-sei=Yu en-aut-mei=Sidney kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= affil-num=1 en-affil= kn-affil=School of Biomedical Sciences, The Chinese University of Hong Kong affil-num=2 en-affil= kn-affil=The Research Core for Interdisciplinary Science, Okayama University affil-num=3 en-affil= kn-affil=Epithelial Cell Biology Research Center, The Chinese University of Hong Kong, affil-num=4 en-affil= kn-affil=School of Biomedical Sciences, The Chinese University of Hong Kong affil-num=5 en-affil= kn-affil=School of Biomedical Sciences, The Chinese University of Hong Kong affil-num=6 en-affil= kn-affil=School of Biomedical Sciences, The Chinese University of Hong Kong affil-num=7 en-affil= kn-affil=Department of Biochemistry, The University of Hong Kong affil-num=8 en-affil= kn-affil=School of Biomedical Sciences, The Chinese University of Hong Kong END start-ver=1.4 cd-journal=joma no-vol=20 cd-vols= no-issue=19 article-no= start-page=4205 end-page=4215 dt-received= dt-revised= dt-accepted= dt-pub-year=2009 dt-pub=20091001 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=mTrs130 Is a Component of a Mammalian TRAPPII Complex, a Rab1 GEF That Binds to COPI-coated Vesicles en-subtitle= kn-subtitle= en-abstract= kn-abstract=The GTPase Rab1 regulates endoplasmic reticulum-Golgi and early Golgi traffic. The guanine nucleotide exchange factor (GEF) or factors that activate Rab1 at these stages of the secretory pathway are currently unknown. Trs130p is a subunit of the yeast TRAPPII (transport protein particle II) complex, a multisubunit tethering complex that is a GEF for the Rab1 homologue Ypt1p. Here, we show that mammalian Trs130 (mTrs130) is a component of an analogous TRAPP complex in mammalian cells, and we describe for the first time the role that this complex plays in membrane traffic. mTRAPPII is enriched on COPI (Coat Protein I)-coated vesicles and buds, but not Golgi cisternae, and it specifically activates Rab1. In addition, we find that mTRAPPII binds to γ1COP, a COPI coat adaptor subunit. The depletion of mTrs130 by short hairpin RNA leads to an increase of vesicles in the vicinity of the Golgi and the accumulation of cargo in an early Golgi compartment. We propose that mTRAPPII is a Rab1 GEF that tethers COPI-coated vesicles to early Golgi membranes. en-copyright= kn-copyright= en-aut-name=YamasakiAkinori en-aut-sei=Yamasaki en-aut-mei=Akinori kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=MenonShekar en-aut-sei=Menon en-aut-mei=Shekar kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=YuSidney en-aut-sei=Yu en-aut-mei=Sidney kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=BarrowmanJemima en-aut-sei=Barrowman en-aut-mei=Jemima kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=MeerlooTimo en-aut-sei=Meerloo en-aut-mei=Timo kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=OorschotViola en-aut-sei=Oorschot en-aut-mei=Viola kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=KlumpermanJudith en-aut-sei=Klumperman en-aut-mei=Judith kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=SatohAyano en-aut-sei=Satoh en-aut-mei=Ayano kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= en-aut-name=Ferro-NovickSusan en-aut-sei=Ferro-Novick en-aut-mei=Susan kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=9 ORCID= affil-num=1 en-affil= kn-affil=Department of Cell Biology, Howard Hughes Medical Institute, Yale University School of Medicine affil-num=2 en-affil= kn-affil=Department of Cell Biology, Howard Hughes Medical Institute, Yale University School of Medicine affil-num=3 en-affil= kn-affil=Department of Cell Biology, Howard Hughes Medical Institute, Yale University School of Medicine affil-num=4 en-affil= kn-affil=Department of Cell Biology, Howard Hughes Medical Institute, Yale University School of Medicine affil-num=5 en-affil= kn-affil=Department of Cellular and Molecular Medicine, University of California at San Diego affil-num=6 en-affil= kn-affil=Cell Microscopy Center, Department of Cell Biology, University Medical Center Utrecht affil-num=7 en-affil= kn-affil=Cell Microscopy Center, Department of Cell Biology, University Medical Center Utrecht affil-num=8 en-affil= kn-affil=Department of Cell Biology, Yale University School of Medicine affil-num=9 en-affil= kn-affil=Department of Cell Biology, Howard Hughes Medical Institute, Yale University School of Medicine END