Single-molecule tracking of PprI in D. radiodurans without interference of autoblinking
Autoblinking is a widespread phenomenon and exhibits high level of intensity in some bacteria. In Deinococcus radiodurans (D. radiodurans), strong autoblinking was found to be indistinguishable from PAmCherry and greatly prevented single-molecule tracking of proteins of interest. Here we employed th...
| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-11-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1256711/full |
| _version_ | 1797641134655143936 |
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| author | Fanfan Zhai Fanfan Zhai Fanfan Zhai Li Hao Xiaomin Chen Ting Jiang Qianhong Guo Qianhong Guo Liping Xie Liping Xie Ying Ma Xiaobo Du Zhiqin Zheng Zhiqin Zheng Zhiqin Zheng Kun Chen Kun Chen Jun Fan Jun Fan |
| author_facet | Fanfan Zhai Fanfan Zhai Fanfan Zhai Li Hao Xiaomin Chen Ting Jiang Qianhong Guo Qianhong Guo Liping Xie Liping Xie Ying Ma Xiaobo Du Zhiqin Zheng Zhiqin Zheng Zhiqin Zheng Kun Chen Kun Chen Jun Fan Jun Fan |
| author_sort | Fanfan Zhai |
| collection | DOAJ |
| description | Autoblinking is a widespread phenomenon and exhibits high level of intensity in some bacteria. In Deinococcus radiodurans (D. radiodurans), strong autoblinking was found to be indistinguishable from PAmCherry and greatly prevented single-molecule tracking of proteins of interest. Here we employed the bright photoswitchable fluorescent protein mMaple3 to label PprI, one essential DNA repair factor, and characterized systematically the fluorescence intensity and bleaching kinetics of both autoblinking and PprI-mMaple3 molecules within cells grown under three different conditions. Under minimal media, we can largely separate autoblinking from mMaple3 molecules and perform reliably single-molecule tracking of PprI in D. radiodurans, by means of applying signal-to-noise ratio and constraining the minimal length for linking the trajectories. We observed three states of PprI molecules, which bear different subcellular localizations and distinct functionalities. Our strategy provides a useful means to study the dynamics and distributions of proteins of interest in bacterial cells with high level of autoblinking. |
| first_indexed | 2024-03-11T13:40:12Z |
| format | Article |
| id | doaj.art-4530496036ee49e9a09c07c68ffbf4e7 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-03-11T13:40:12Z |
| publishDate | 2023-11-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-4530496036ee49e9a09c07c68ffbf4e72023-11-02T13:02:36ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2023-11-011410.3389/fmicb.2023.12567111256711Single-molecule tracking of PprI in D. radiodurans without interference of autoblinkingFanfan Zhai0Fanfan Zhai1Fanfan Zhai2Li Hao3Xiaomin Chen4Ting Jiang5Qianhong Guo6Qianhong Guo7Liping Xie8Liping Xie9Ying Ma10Xiaobo Du11Zhiqin Zheng12Zhiqin Zheng13Zhiqin Zheng14Kun Chen15Kun Chen16Jun Fan17Jun Fan18School of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, ChinaInstitute of Fundamental and Frontier Sciences, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaNHC Key Laboratory of Nuclear Technology Medical Transformation (Mianyang Central Hospital), Mianyang, Sichuan, ChinaInstitute of Fundamental and Frontier Sciences, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaInstitute of Fundamental and Frontier Sciences, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaInstitute of Fundamental and Frontier Sciences, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaSchool of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, ChinaInstitute of Fundamental and Frontier Sciences, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaSchool of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, ChinaInstitute of Fundamental and Frontier Sciences, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaNHC Key Laboratory of Nuclear Technology Medical Transformation (Mianyang Central Hospital), Mianyang, Sichuan, ChinaNHC Key Laboratory of Nuclear Technology Medical Transformation (Mianyang Central Hospital), Mianyang, Sichuan, ChinaSchool of Life Sciences and Engineering, Southwest University of Science and Technology, Mianyang, Sichuan, ChinaNHC Key Laboratory of Nuclear Technology Medical Transformation (Mianyang Central Hospital), Mianyang, Sichuan, ChinaSchool of Biological Engineering and Wuliangye Liquor, Sichuan University of Science and Engineering, Yibin, Sichuan, ChinaSchool of Optoelectronic Science and Engineering, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaYangtze Delta Region Institute (Huzhou), University of Electronic Science and Technology of China, Huzhou, Zhejiang, ChinaInstitute of Fundamental and Frontier Sciences, University of Electronic Science and Technology of China, Chengdu, Sichuan, ChinaYangtze Delta Region Institute (Huzhou), University of Electronic Science and Technology of China, Huzhou, Zhejiang, ChinaAutoblinking is a widespread phenomenon and exhibits high level of intensity in some bacteria. In Deinococcus radiodurans (D. radiodurans), strong autoblinking was found to be indistinguishable from PAmCherry and greatly prevented single-molecule tracking of proteins of interest. Here we employed the bright photoswitchable fluorescent protein mMaple3 to label PprI, one essential DNA repair factor, and characterized systematically the fluorescence intensity and bleaching kinetics of both autoblinking and PprI-mMaple3 molecules within cells grown under three different conditions. Under minimal media, we can largely separate autoblinking from mMaple3 molecules and perform reliably single-molecule tracking of PprI in D. radiodurans, by means of applying signal-to-noise ratio and constraining the minimal length for linking the trajectories. We observed three states of PprI molecules, which bear different subcellular localizations and distinct functionalities. Our strategy provides a useful means to study the dynamics and distributions of proteins of interest in bacterial cells with high level of autoblinking.https://www.frontiersin.org/articles/10.3389/fmicb.2023.1256711/fullsingle-molecule trackingPprI proteinmMaple3autoblinkingDeinococcus radiodurans |
| spellingShingle | Fanfan Zhai Fanfan Zhai Fanfan Zhai Li Hao Xiaomin Chen Ting Jiang Qianhong Guo Qianhong Guo Liping Xie Liping Xie Ying Ma Xiaobo Du Zhiqin Zheng Zhiqin Zheng Zhiqin Zheng Kun Chen Kun Chen Jun Fan Jun Fan Single-molecule tracking of PprI in D. radiodurans without interference of autoblinking Frontiers in Microbiology single-molecule tracking PprI protein mMaple3 autoblinking Deinococcus radiodurans |
| title | Single-molecule tracking of PprI in D. radiodurans without interference of autoblinking |
| title_full | Single-molecule tracking of PprI in D. radiodurans without interference of autoblinking |
| title_fullStr | Single-molecule tracking of PprI in D. radiodurans without interference of autoblinking |
| title_full_unstemmed | Single-molecule tracking of PprI in D. radiodurans without interference of autoblinking |
| title_short | Single-molecule tracking of PprI in D. radiodurans without interference of autoblinking |
| title_sort | single molecule tracking of ppri in d radiodurans without interference of autoblinking |
| topic | single-molecule tracking PprI protein mMaple3 autoblinking Deinococcus radiodurans |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1256711/full |
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