title: Facile manipulation of protein localization in fission yeast through binding of GFP-binding protein to GFP.
abstract: GFP-binding protein (or GBP) has been recently developed in various systems and organisms as an efficient tool to purify GFP-fusion proteins. Due to the high affinity between GBP and GFP or GFP variants, this GBP-based approach is also ideally suited to alter the localization of functional proteins in live cells. In order to facilitate the wide use of the GBP-targeting approach in the fission yeastSchizosaccharomyces pombe, we developed a set of pFA6a-, pJK148- and pUC119-based vectors containing GBP- or GBP-mCherry-coding sequences and variants of induciblenmt1or constitutiveadh1promoters that result in different levels of expression. The GBP or GBP-mCherry fragments can serve as cassettes for N- or C-terminal genomic tagging of genes of interest. We illustrated the application of these vectors in the construction of yeast strains with Dma1 or Cdc7 tagged with GBP-mCherry and efficient targeting of Dma1- or Cdc7-GBP-mCherry to the spindle pole body by Sid4-GFP. This series of vectors should help to facilitate the application of the GBP-targeting approach in manipulating protein localization and the analysis of gene function in fission yeast, at the level of single genes, as well as at a systematic scale.
authors.affiliation: State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China.
authors.forename: Ying-Hui
authors.identifier: 0000-0002-2464-7621
authors.initials: YH
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China.
authors.forename: Gao-Yuan
authors.identifier: 0000-0003-3165-0818
authors.initials: GY
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China.
authors.forename: Hao-Chao
authors.identifier: 0000-0003-2992-5026
authors.initials: HC
authors.lastname: Hao
authors.affiliation: State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China.
authors.forename: Chun-Jiang
authors.identifier: 0000-0001-7374-6063
authors.initials: CJ
authors.lastname: Chao
authors.affiliation: State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China jinquanwen@xmu.edu.cn wangyamei@xmu.edu.cn.
authors.forename: Yamei
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Cellular Stress Biology, School of Life Sciences, Xiamen University, Xiamen, Fujian 361102, China jinquanwen@xmu.edu.cn wangyamei@xmu.edu.cn.
authors.forename: Quan-Wen
authors.identifier: 0000-0001-6146-6910
authors.initials: QW
authors.lastname: Jin
mesh_terms: D001483:Base Sequence; D003584:Cytological Techniques / Q000379:methods*; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D008156:Luciferases / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012568:Schizosaccharomyces / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000378:metabolism; D064048:Spindle Pole Bodies / Q000378:metabolism
chemical_list: D011993:Recombinant Fusion Proteins; D029702:Schizosaccharomyces pombe Proteins; D049452:Green Fluorescent Proteins; D008156:Luciferases
keywords: Adh; Fission yeast; GBP; GBP–GFP high affinity; GBP–mCherry; Nmt; Targeted protein localization