Cell Structure and Function
Online ISSN : 1347-3700
Print ISSN : 0386-7196
ISSN-L : 0386-7196
Folding Latency of Fluorescent Proteins Affects the Mitochondrial Localization of Fusion Proteins
Sayaka KashiwagiYoichiro FujiokaAya O. SatohAiko YoshidaMari FujiokaPrabha NepalAtsushi TsuzukiOzora AokiSarad PaudelHitoshi SasajimaYusuke Ohba
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Supplementary material

2019 Volume 44 Issue 2 Pages 183-194

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Abstract

The discovery of fluorescent proteins (FPs) has revolutionized cell biology. The fusion of targeting sequences to FPs enables the investigation of cellular organelles and their dynamics; however, occasionally, such fluorescent fusion proteins (FFPs) exhibit behavior different from that of the native proteins. Here, we constructed a color pallet comprising different organelle markers and found that FFPs targeted to the mitochondria were mislocalized when fused to certain types of FPs. Such FPs included several variants of Aequorea victoria green FP (avGFP) and a monomeric variant of the red FP. Because the FFPs that are mislocalized include FPs with faster maturing or folding mutations, the increase in the maturation rate is likely to prevent their expected localization. Indeed, when we reintroduced amino acid substitutions so that the FP sequences were equivalent to that of wild-type avGFP, FFP localization to the mitochondria was significantly enhanced. Moreover, similar amino acid substitutions improved the localization of mitochondria-targeted pHluorin, which is a pH-sensitive variant of GFP, and its capability to monitor pH changes in the mitochondrial matrix. Our findings demonstrate the importance of selecting FPs that maximize FFP function.

Key words: fluorescent protein, organelle, fusion protein, mitochondria

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© 2019 The Author(s) CC-BY 4.0 (Submission before October 2016: Copyright © Japan Society for Cell Biology)

Copyright: ©2019 The Author(s). This is an open access article distributed under the terms of the Creative Commons BY (Attribution) License (https://creativecommons.org/licenses/by/4.0/legalcode), which permits the unrestricted distribution, reproduction and use of the article provided the original source and authors are credited.
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