Development and application of a recombination-based library versus library highthroughput yeast two-hybrid (RLL-Y2H) screening system

Fang Yang; Yingying Lei; Meiling Zhou; Qili Yao; Yichao Han; Xiang Wu; Wanshun Zhong; Chenghang Zhu; Weize Xu; Ran Tao; Xi Chen; Da Lin; Khaista Rahman; Rohit Tyagi; Zeshan Habib; Shaobo Xiao; Dang Wang; Yang Yu; Huanchun Chen; Zhenfang Fu; Gang Cao

doi:10.1093/nar/gkx1173

Development and application of a recombination-based library versus library highthroughput yeast two-hybrid (RLL-Y2H) screening system

Fang Yang, Yingying Lei, Meiling Zhou, Qili Yao, Yichao Han, Xiang Wu, Wanshun Zhong, Chenghang Zhu, Weize Xu, Ran Tao, Xi Chen, Da Lin, Khaista Rahman, Rohit Tyagi, Zeshan Habib, Shaobo Xiao, Dang Wang, Yang Yu, Huanchun Chen, Zhenfang FuGang Cao^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

31 Citations (Scopus)

Abstract

Protein-protein interaction (PPI) network maintains proper function of all organisms. Simple highthroughput technologies are desperately needed to delineate the landscape of PPI networks. While recent state-of-the-art yeast two-hybrid (Y2H) systems improved screening efficiency, either individual colony isolation, library preparation arrays, gene barcoding or massive sequencing are still required. Here, we developed a recombination-based 'library vs library' Y2H system (RLL-Y2H), by which multilibrary screening can be accomplished in a single pool without any individual treatment. This system is based on the phiC31 integrase-mediated integration between bait and prey plasmids. The integrated fragments were digested by MmeIand subjected to deep sequencing to decode the interaction matrix. We applied this system to decipher the trans-kingdom interactome betweenMycobacterium tuberculosisand host cells and further identified Rv2427c interfering with the phagosome-lysosome fusion. This concept can also be applied to other systems to screen protein-RNA and protein-DNA interactions and delineate signaling landscape in cells.

Original language	English
Article number	e17
Journal	Nucleic Acids Research
Volume	46
Issue number	3
DOIs	https://doi.org/10.1093/nar/gkx1173
Publication status	Published - 16 Feb 2018
Externally published	Yes

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Yang, F., Lei, Y., Zhou, M., Yao, Q., Han, Y., Wu, X., Zhong, W., Zhu, C., Xu, W., Tao, R., Chen, X., Lin, D., Rahman, K., Tyagi, R., Habib, Z., Xiao, S., Wang, D., Yu, Y., Chen, H., ... Cao, G. (2018). Development and application of a recombination-based library versus library highthroughput yeast two-hybrid (RLL-Y2H) screening system. Nucleic Acids Research, 46(3), Article e17. https://doi.org/10.1093/nar/gkx1173

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title = "Development and application of a recombination-based library versus library highthroughput yeast two-hybrid (RLL-Y2H) screening system",

abstract = "Protein-protein interaction (PPI) network maintains proper function of all organisms. Simple highthroughput technologies are desperately needed to delineate the landscape of PPI networks. While recent state-of-the-art yeast two-hybrid (Y2H) systems improved screening efficiency, either individual colony isolation, library preparation arrays, gene barcoding or massive sequencing are still required. Here, we developed a recombination-based 'library vs library' Y2H system (RLL-Y2H), by which multilibrary screening can be accomplished in a single pool without any individual treatment. This system is based on the phiC31 integrase-mediated integration between bait and prey plasmids. The integrated fragments were digested by MmeIand subjected to deep sequencing to decode the interaction matrix. We applied this system to decipher the trans-kingdom interactome betweenMycobacterium tuberculosisand host cells and further identified Rv2427c interfering with the phagosome-lysosome fusion. This concept can also be applied to other systems to screen protein-RNA and protein-DNA interactions and delineate signaling landscape in cells.",

author = "Fang Yang and Yingying Lei and Meiling Zhou and Qili Yao and Yichao Han and Xiang Wu and Wanshun Zhong and Chenghang Zhu and Weize Xu and Ran Tao and Xi Chen and Da Lin and Khaista Rahman and Rohit Tyagi and Zeshan Habib and Shaobo Xiao and Dang Wang and Yang Yu and Huanchun Chen and Zhenfang Fu and Gang Cao",

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Yang, F, Lei, Y, Zhou, M, Yao, Q, Han, Y, Wu, X, Zhong, W, Zhu, C, Xu, W, Tao, R, Chen, X, Lin, D, Rahman, K, Tyagi, R, Habib, Z, Xiao, S, Wang, D, Yu, Y, Chen, H, Fu, Z & Cao, G 2018, 'Development and application of a recombination-based library versus library highthroughput yeast two-hybrid (RLL-Y2H) screening system', Nucleic Acids Research, vol. 46, no. 3, e17. https://doi.org/10.1093/nar/gkx1173

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T1 - Development and application of a recombination-based library versus library highthroughput yeast two-hybrid (RLL-Y2H) screening system

AU - Yang, Fang

AU - Lei, Yingying

AU - Zhou, Meiling

AU - Yao, Qili

AU - Han, Yichao

AU - Wu, Xiang

AU - Zhong, Wanshun

AU - Zhu, Chenghang

AU - Xu, Weize

AU - Tao, Ran

AU - Chen, Xi

AU - Lin, Da

AU - Rahman, Khaista

AU - Tyagi, Rohit

AU - Habib, Zeshan

AU - Xiao, Shaobo

AU - Wang, Dang

AU - Yu, Yang

AU - Chen, Huanchun

AU - Fu, Zhenfang

AU - Cao, Gang

PY - 2018/2/16

Y1 - 2018/2/16

N2 - Protein-protein interaction (PPI) network maintains proper function of all organisms. Simple highthroughput technologies are desperately needed to delineate the landscape of PPI networks. While recent state-of-the-art yeast two-hybrid (Y2H) systems improved screening efficiency, either individual colony isolation, library preparation arrays, gene barcoding or massive sequencing are still required. Here, we developed a recombination-based 'library vs library' Y2H system (RLL-Y2H), by which multilibrary screening can be accomplished in a single pool without any individual treatment. This system is based on the phiC31 integrase-mediated integration between bait and prey plasmids. The integrated fragments were digested by MmeIand subjected to deep sequencing to decode the interaction matrix. We applied this system to decipher the trans-kingdom interactome betweenMycobacterium tuberculosisand host cells and further identified Rv2427c interfering with the phagosome-lysosome fusion. This concept can also be applied to other systems to screen protein-RNA and protein-DNA interactions and delineate signaling landscape in cells.

AB - Protein-protein interaction (PPI) network maintains proper function of all organisms. Simple highthroughput technologies are desperately needed to delineate the landscape of PPI networks. While recent state-of-the-art yeast two-hybrid (Y2H) systems improved screening efficiency, either individual colony isolation, library preparation arrays, gene barcoding or massive sequencing are still required. Here, we developed a recombination-based 'library vs library' Y2H system (RLL-Y2H), by which multilibrary screening can be accomplished in a single pool without any individual treatment. This system is based on the phiC31 integrase-mediated integration between bait and prey plasmids. The integrated fragments were digested by MmeIand subjected to deep sequencing to decode the interaction matrix. We applied this system to decipher the trans-kingdom interactome betweenMycobacterium tuberculosisand host cells and further identified Rv2427c interfering with the phagosome-lysosome fusion. This concept can also be applied to other systems to screen protein-RNA and protein-DNA interactions and delineate signaling landscape in cells.

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JO - Nucleic Acids Research

JF - Nucleic Acids Research

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ER -

Development and application of a recombination-based library versus library highthroughput yeast two-hybrid (RLL-Y2H) screening system

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