title: Plastoglobule-Targeting Competence of a Putative Transit Peptide Sequence from Rice Phytoene Synthase 2 in Plastids.
abstract: Plastoglobules (PGs) are thylakoid membrane microdomains within plastids that are known as specialized locations of carotenogenesis. Three rice phytoene synthase proteins (OsPSYs) involved in carotenoid biosynthesis have been identified. Here, the N-terminal 80-amino-acid portion of OsPSY2 (PTp) was demonstrated to be a chloroplast-targeting peptide by displaying cytosolic localization of OsPSY2(ΔPTp):mCherry in rice protoplast, in contrast to chloroplast localization of OsPSY2:mCherry in a punctate pattern. The peptide sequence of a PTp was predicted to harbor two transmembrane domains eligible for a putative PG-targeting signal. To assess and enhance the PG-targeting ability of PTp, the original PTp DNA sequence (PTp) was modified to a synthetic DNA sequence (stPTp), which had 84.4% similarity to the original sequence. The motivation of this modification was to reduce the GC ratio from 75% to 65% and to disentangle the hairpin loop structures ofPTp. These two DNA sequences were fused to the sequence of the synthetic green fluorescent protein (sGFP) and drove GFP expression with different efficiencies. In particular, the RNA and protein levels ofstPTp-sGFPwere slightly improved to 1.4-fold and 1.3-fold more than those of sGFP, respectively. The green fluorescent signals of their mature proteins were all observed as speckle-like patterns with slightly blurred stromal signals in chloroplasts. These discrete green speckles of PTp-sGFP and stPTp-sGFP corresponded exactly to the red fluorescent signal displayed by OsPSY2:mCherry in both etiolated and greening protoplasts and it is presumed to correspond to distinct PGs. In conclusion, we identified PTp as a transit peptide sequence facilitating preferential translocation of foreign proteins to PGs, and developed an improvedPTpsequence, a stPTp, which is expected to be very useful for applications in plant biotechnologies requiring precise micro-compartmental localization in plastids.
authors.affiliation: Department of Genetic Engineering and Graduate School of Biotechnology, College of Life Sciences, Kyung Hee University, Yongin-si 17104, Korea. minkyou@khu.ac.kr.|Crop Biotech Institute, Kyung Hee University, Yongin-si 17104, Korea. minkyou@khu.ac.kr.
authors.forename: Min Kyoung
authors.initials: MK
authors.lastname: You
authors.affiliation: Crop Biotech Institute, Kyung Hee University, Yongin-si 17104, Korea. flowerjin@nate.com.
authors.forename: Jin Hwa
authors.initials: JH
authors.lastname: Kim
authors.affiliation: Department of Genetic Engineering and Graduate School of Biotechnology, College of Life Sciences, Kyung Hee University, Yongin-si 17104, Korea. yeojin@khu.ac.kr.|Crop Biotech Institute, Kyung Hee University, Yongin-si 17104, Korea. yeojin@khu.ac.kr.
authors.forename: Yeo Jin
authors.initials: YJ
authors.lastname: Lee
authors.affiliation: Crop Biotech Institute, Kyung Hee University, Yongin-si 17104, Korea. yesol@khu.ac.kr.
authors.forename: Ye Sol
authors.initials: YS
authors.lastname: Jeong
authors.affiliation: Department of Genetic Engineering and Graduate School of Biotechnology, College of Life Sciences, Kyung Hee University, Yongin-si 17104, Korea. sunhwa@khu.ac.kr.|Crop Biotech Institute, Kyung Hee University, Yongin-si 17104, Korea. sunhwa@khu.ac.kr.
authors.forename: Sun-Hwa
authors.initials: SH
authors.lastname: Ha
mesh_terms: D001482:Base Composition; D060365:Chloroplast Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence; D051232:Geranylgeranyl-Diphosphate Geranylgeranyltransferase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012275:Oryza / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D021382:Protein Sorting Signals / Q000235:genetics; D021381:Protein Transport; D020524:Thylakoids / Q000378:metabolism*
chemical_list: D060365:Chloroplast Proteins; D021382:Protein Sorting Signals; C078737:chloroplast transit peptides; D051232:Geranylgeranyl-Diphosphate Geranylgeranyltransferase
keywords: phytoene synthase; plastid; plastoglobule; protoplast; rice; transit peptide