九州大学-研究者情報 [栁谷耕太 (准教授) 生体防御医学研究所細胞機能制御学部門]

Kota Yanagitani, Kenji Kohno, Nascent chain-mediated localization of mRNA on the endoplasmic reticulum as an important step of unfolded protein response.
Regulatory Nascent Polypeptides, (eds., Ito K), pp.291-310
, Springer Japan, 10.1007/978-4-431-55052-5, pp.291-310, 2014.01, A system for homeostatic control of proteome, namely proteostasis, in a cell is conserved through a wide range of living organisms. The system is composed of a sensor for misfolded proteins that arise from perturbed proteostasis and signaling machinery that induces the expression of genes for molecular chaperones and proteolytic factors to counterbalance the proteostasis perturbation. In eukaryotes, distinct cellular response mechanisms have been elaborated for perturbed proteostasis within each organelle. In these cases, a sensor molecule recognizes misfolded proteins in the organelle and somehow transmits the signal to the nucleus across the organelle membrane. In the case of the endoplasmic reticulum (ER),
proteostatic perturbation is sensed by an ER-resident transmembrane endoribonuclease inositol requiring 1 (IRE1) that catalyzes unconventional splicing of a precursor mRNA on the ER membrane. The resulting spliced form of the mRNA encodes a transcription factor that enters the nucleus and evokes a cellular response to counterbalance the perturbed proteostasis by activating transcription of target genes. In this chapter, we discuss the recently characterized mechanism, in which a nascent polypeptide encoded by the splicing-substrate mRNA directs localization of its own mRNA onto the ER membrane as an essential step for the efficient unconventional splicing of the substrate mRNA upon ER stresses. In particular, we focus on the mechanism that targets the unspliced precursor form of X-box binding protein 1 (XBP1u) mRNA to the ER membrane in metazoan cells. Importantly, translational pausing plays pivotal roles that ensure the nascent chain-mediated ER-targeting of XBP1u mRNA. We also discuss the prerequisites and generality of nascent chain-mediated mRNA localization in the cell..

主要原著論文

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1.	Yanagitani K, Imagawa Y, Iwawaki T, Hosoda A, Saito M, Kimata Y, Kohno K, Cotranslational targeting of XBP1 protein to the membrane promotes cytoplasmic splicing of its own mRNA, Mol Cell, 34, 2, 191-200, 2009.04, Endoplasmic reticulum (ER) stress triggers the cytoplasmic splicing of XBP1 mRNA by the transmembrane endoribonuclease IRE1alpha, resulting in activation of the unfolded protein response, which maintains ER homeostasis. We show that the unspliced XBP1 (XBP1u) mRNA is localized to the membrane, although its product is neither a secretory nor a membrane protein and is released to the cytosol after splicing. Biochemical and mutagenic analyses demonstrated that membrane localization of XBP1u mRNA required its in-frame translation. An insertional frame-shift mutation greatly diminished both membrane localization and splicing of the XBP1u mRNA. Furthermore, membrane localization was compromised by puromycin treatment and required a hydrophobic region within XBP1u. These data demonstrate that the nascent XBP1u polypeptide recruits its own mRNA to the membrane. This system serves to enhance cytoplasmic splicing and could facilitate a more rapid response to ER stress, and represents a unique way of cotranslational protein targeting coupled to mRNA maturation..
2.	Yanagitani K, Kimata Y, Kadokura H, Kohno K., Translational pausing ensures membrane targeting and cytoplasmic splicing of XBP1u mRNA., Science, 331, 6017, 586-589, 2011.02, Upon endoplasmic reticulum (ER) stress, an endoribonuclease, inositol-requiring enzyme-1α, splices the precursor unspliced form of X-box-binding protein 1 messenger RNA (XBP1u mRNA) on the ER membrane to yield an active transcription factor (XBP1s), leading to the alleviation of the stress. The nascent peptide encoded by XBP1u mRNA drags the mRNA-ribosome-nascent chain (R-RNC) complex to the membrane for efficient cytoplasmic splicing. We found that translation of the XBP1u mRNA was briefly paused to stabilize the R-RNC complex. Mutational analysis of XBP1u revealed an evolutionarily conserved peptide module at the carboxyl terminus that was responsible for the translational pausing and was required for the efficient targeting and splicing of the XBP1u mRNA. Thus, translational pausing may be used for unexpectedly diverse cellular processes in mammalian cells..
3.	Kanda S, Yanagitani K (共筆頭著, 共責任著者), Yokota Y, Esaki Y, Kohno K, Autonomous translational pausing is required for XBP1u mRNA recruitment to the ER via the SRP pathway., Proc Natl Acad Sci USA, 113, 40, E5886-E5895, 2016.10, Unconventional mRNA splicing on the endoplasmic reticulum (ER) membrane is the sole conserved mechanism in eukaryotes to transmit information regarding misfolded protein accumulation to the nucleus to activate the stress response. In metazoans, the unspliced form of X-box-binding protein 1 (XBP1u) mRNA is recruited to membranes as a ribosome nascent chain (RNC) complex for efficient splicing. We previously reported that both hydrophobic (HR2) and translational pausing regions of XBP1u are important for the recruitment of its own mRNA to membranes. However, its precise location and the molecular mechanism of translocation are unclear. We show that XBP1u-RNC is specifically recruited to the ER membrane in an HR2- and translational pausing-dependent manner by immunostaining, fluorescent recovery after photobleaching, and biochemical analyses. Notably, translational pausing during XBP1u synthesis is indispensable for the recognition of HR2 by the signal recognition particle (SRP), resulting in efficient ER-specific targeting of the complex, similar to secretory protein targeting to the ER. On the ER, the XBP1u nascent chain is transferred from the SRP to the translocon; however, it cannot pass through the translocon or insert into the membrane. Therefore, our results support a noncanonical mechanism by which mRNA substrates are recruited to the ER for unconventional splicing..
4.	Yanagitani K, Juszkiewicz S, Hegde RS., UBE2O is a quality control factor for orphans of multiprotein complexes., Science, doi: 10.1126/science.aan0178, 357, 6350, 472-475, 2017.08, Many nascent proteins are assembled into multiprotein complexes of defined stoichiometry. Imbalances in the synthesis of individual subunits result in orphans. How orphans are selectively eliminated to maintain protein homeostasis is poorly understood. Here, we found that the conserved ubiquitin-conjugating enzyme UBE2O directly recognized juxtaposed basic and hydrophobic patches on unassembled proteins to mediate ubiquitination without a separate ubiquitin ligase. In reticulocytes, where UBE2O is highly up-regulated, unassembled α-globin molecules that failed to assemble with β-globin were selectively ubiquitinated by UBE2O. In nonreticulocytes, ribosomal proteins that did not engage nuclear import factors were targets for UBE2O. Thus, UBE2O is a self-contained quality control factor that comprises substrate recognition and ubiquitin transfer activities within a single protein to efficiently target orphans of multiprotein complexes for degradation..

主要学会発表等

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学会活動

所属学会名

日本分子生物学会

日本細胞生物学会

その他の研究活動

海外渡航状況, 海外での教育研究歴

MRC Laboratory of Molecular Biology, UnitedKingdom, 2013.02～2016.03.

受賞

第9回梅園賞, 奈良先端科学技術大学院大学, 2012.08.

第28回井上研究奨励賞, 井上科学振興財団, 2012.02.

第63回日本細胞生物学会・若手優秀発表賞, 日本細胞生物学会, 2011.06.

全体班会議・優秀ポスター賞, 特定領域研究“タンパク質の社会, 2009.11.

博士後期過程・最優秀学生賞, 奈良先端科学技術大学院大学, 2009.03.

全体班会議・最優秀ポスター賞 , 特定領域研究“タンパク質の社会, 2008.11.

バイオCOEサマーキャンプ・優秀発表賞, 奈良先端科学技術大学院大学, 2006.09.

バイオCOEサマーキャンプ・優秀発表賞, 奈良先端科学技術大学院大学, 2005.09.

研究資金

科学研究費補助金の採択状況(文部科学省、日本学術振興会)

2019年度～2021年度, 基盤研究(C), 代表, オルガネラ量の恒常性を保つシステムの根底原理の解明.

日本学術振興会への採択状況(科学研究費補助金以外)

2012年度～2015年度, 海外特別研究員, 代表, トランスロコンの閉塞に対する細胞応答機構の探索.

競争的資金(受託研究を含む)の採択状況

2011年度～2012年度, 科学研究費補助金 (文部科学省）, 代表, オルガネラノックダウン法の開発を契機としたオルガネラ量補償機構の解析.

2019年度～2021年度, 科学研究費補助金 (文部科学省）, 代表, オルガネラ量の恒常性を保つシステムの根底原理の解明.

九大関連コンテンツ

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生体防御医学研究所


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