An unexpected role for leucyl aminopeptidase in UV tolerance revealed by a genome-wide fitness assessment in a model cyanobacterium

Elliot L. Weiss; Mingxu Fang; Arnaud Taton; Richard Szubin; Bernhard Palsson; B. Greg Mitchell; Susan S. Golden

doi:10.1073/pnas.2211789119

An unexpected role for leucyl aminopeptidase in UV tolerance revealed by a genome-wide fitness assessment in a model cyanobacterium

Elliot L. Weiss
, Mingxu Fang
, Arnaud Taton
, Richard Szubin
, Bernhard Palsson
, B. Greg Mitchell
, Susan S. Golden

University of Iceland

University of Iceland

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

Abstract

UV radiation (UVR) has significant physiological effects on organisms living at or near the Earth's surface, yet the full suite of genes required for fitness of a photosynthetic organism in a UVR-rich environment remains unknown. This study reports a genome-wide fitness assessment of the genes that affect UVR tolerance under environmentally relevant UVR dosages in the model cyanobacterium Synechococcus elongatus PCC 7942. Our results highlight the importance of specific genes that encode proteins involved in DNA repair, glutathione synthesis, and the assembly and maintenance of photosystem II, as well as genes that encode hypothetical proteins and others without an obvious connection to canonical methods of UVR tolerance. Disruption of a gene that encodes a leucyl aminopeptidase (LAP) conferred the greatest UVR-specific decrease in fitness. Enzymatic assays demonstrated a strong pH-dependent affinity of the LAP for the dipeptide cysteinyl-glycine, suggesting an involvement in glutathione catabolism as a function of night-time cytosolic pH level. A low differential expression of the LAP gene under acute UVR exposure suggests that its relative importance would be overlooked in transcript-dependent screens. Subsequent experiments revealed a similar UVR-sensitivity phenotype in LAP knockouts of other organisms, indicating conservation of the functional role of LAPs in UVR tolerance.

Original language	English
Article number	e2211789119
Pages (from-to)	e2211789119
Journal	Proceedings of the National Academy of Sciences of the United States of America
Volume	119
Issue number	45
DOIs	https://doi.org/10.1073/pnas.2211789119
Publication status	Published - 8 Nov 2022

Bibliographical note

Funding Information: We thank David Welkie and Benjamin Rubin for consultation on experimental design, Emily Pierce for thoughtful comments on the manuscript, and Ryan Simkovsky for providing the assembled plasmid pAM5217. This work was funded by the National Science Foundation through the University of California, San Diego Materials Research Science and Engineering Center (DMR-2011924) and Graduate Research Fellowship Program (GRFP) DGE-1650112 (to E.L.W.). Support for RNA sequencing was provided by the YC Fung Chair in Bioengineering (to B.Ø.P.). M.F., A.T., and S.S.G. were supported by the National Institute of General Medical Sciences of the NIH under award R35GM118290. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. Funding Information: ACKNOWLEDGMENTS. We thank David Welkie and Benjamin Rubin for consultation on experimental design, Emily Pierce for thoughtful comments on the manuscript, and Ryan Simkovsky for providing the assembled plasmid pAM5217. This work was funded by the National Science Foundation through the University of California, San Diego Materials Research Science and Engineering Center (DMR-2011924) and Graduate Research Fellowship Program (GRFP) DGE-1650112 (to E.L.W.). Support for RNA sequencing was provided by the YC Fung Chair in Bioengineering (to B.Ø.P.). M.F., A.T., and S.S.G. were supported by the National Institute of General Medical Sciences of the NIH under award R35GM118290. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. Publisher Copyright: Copyright © 2022 the Author(s).

Other keywords

RB-TnSeq
UV radiation
cyanobacteria
fitness
leucyl aminopeptidase

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10.1073/pnas.2211789119

Cite this

Weiss, E. L., Fang, M., Taton, A., Szubin, R., Palsson, B., Mitchell, B. G., & Golden, S. S. (2022). An unexpected role for leucyl aminopeptidase in UV tolerance revealed by a genome-wide fitness assessment in a model cyanobacterium. Proceedings of the National Academy of Sciences of the United States of America, 119(45), e2211789119. Article e2211789119. https://doi.org/10.1073/pnas.2211789119

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title = "An unexpected role for leucyl aminopeptidase in UV tolerance revealed by a genome-wide fitness assessment in a model cyanobacterium",

abstract = "UV radiation (UVR) has significant physiological effects on organisms living at or near the Earth's surface, yet the full suite of genes required for fitness of a photosynthetic organism in a UVR-rich environment remains unknown. This study reports a genome-wide fitness assessment of the genes that affect UVR tolerance under environmentally relevant UVR dosages in the model cyanobacterium Synechococcus elongatus PCC 7942. Our results highlight the importance of specific genes that encode proteins involved in DNA repair, glutathione synthesis, and the assembly and maintenance of photosystem II, as well as genes that encode hypothetical proteins and others without an obvious connection to canonical methods of UVR tolerance. Disruption of a gene that encodes a leucyl aminopeptidase (LAP) conferred the greatest UVR-specific decrease in fitness. Enzymatic assays demonstrated a strong pH-dependent affinity of the LAP for the dipeptide cysteinyl-glycine, suggesting an involvement in glutathione catabolism as a function of night-time cytosolic pH level. A low differential expression of the LAP gene under acute UVR exposure suggests that its relative importance would be overlooked in transcript-dependent screens. Subsequent experiments revealed a similar UVR-sensitivity phenotype in LAP knockouts of other organisms, indicating conservation of the functional role of LAPs in UVR tolerance.",

keywords = "RB-TnSeq, UV radiation, cyanobacteria, fitness, leucyl aminopeptidase",

author = "Weiss, \{Elliot L.\} and Mingxu Fang and Arnaud Taton and Richard Szubin and Bernhard Palsson and Mitchell, \{B. Greg\} and Golden, \{Susan S.\}",

note = "Funding Information: We thank David Welkie and Benjamin Rubin for consultation on experimental design, Emily Pierce for thoughtful comments on the manuscript, and Ryan Simkovsky for providing the assembled plasmid pAM5217. This work was funded by the National Science Foundation through the University of California, San Diego Materials Research Science and Engineering Center (DMR-2011924) and Graduate Research Fellowship Program (GRFP) DGE-1650112 (to E.L.W.). Support for RNA sequencing was provided by the YC Fung Chair in Bioengineering (to B.{\O}.P.). M.F., A.T., and S.S.G. were supported by the National Institute of General Medical Sciences of the NIH under award R35GM118290. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. Funding Information: ACKNOWLEDGMENTS. We thank David Welkie and Benjamin Rubin for consultation on experimental design, Emily Pierce for thoughtful comments on the manuscript, and Ryan Simkovsky for providing the assembled plasmid pAM5217. This work was funded by the National Science Foundation through the University of California, San Diego Materials Research Science and Engineering Center (DMR-2011924) and Graduate Research Fellowship Program (GRFP) DGE-1650112 (to E.L.W.). Support for RNA sequencing was provided by the YC Fung Chair in Bioengineering (to B.{\O}.P.). M.F., A.T., and S.S.G. were supported by the National Institute of General Medical Sciences of the NIH under award R35GM118290. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. Publisher Copyright: Copyright {\textcopyright} 2022 the Author(s).",

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Weiss, EL, Fang, M, Taton, A, Szubin, R, Palsson, B, Mitchell, BG & Golden, SS 2022, 'An unexpected role for leucyl aminopeptidase in UV tolerance revealed by a genome-wide fitness assessment in a model cyanobacterium', Proceedings of the National Academy of Sciences of the United States of America, vol. 119, no. 45, e2211789119, pp. e2211789119. https://doi.org/10.1073/pnas.2211789119

An unexpected role for leucyl aminopeptidase in UV tolerance revealed by a genome-wide fitness assessment in a model cyanobacterium. / Weiss, Elliot L.; Fang, Mingxu; Taton, Arnaud et al.
In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 119, No. 45, e2211789119, 08.11.2022, p. e2211789119.

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

TY - JOUR

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AU - Fang, Mingxu

AU - Taton, Arnaud

AU - Szubin, Richard

AU - Palsson, Bernhard

AU - Mitchell, B. Greg

AU - Golden, Susan S.

N1 - Funding Information: We thank David Welkie and Benjamin Rubin for consultation on experimental design, Emily Pierce for thoughtful comments on the manuscript, and Ryan Simkovsky for providing the assembled plasmid pAM5217. This work was funded by the National Science Foundation through the University of California, San Diego Materials Research Science and Engineering Center (DMR-2011924) and Graduate Research Fellowship Program (GRFP) DGE-1650112 (to E.L.W.). Support for RNA sequencing was provided by the YC Fung Chair in Bioengineering (to B.Ø.P.). M.F., A.T., and S.S.G. were supported by the National Institute of General Medical Sciences of the NIH under award R35GM118290. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. Funding Information: ACKNOWLEDGMENTS. We thank David Welkie and Benjamin Rubin for consultation on experimental design, Emily Pierce for thoughtful comments on the manuscript, and Ryan Simkovsky for providing the assembled plasmid pAM5217. This work was funded by the National Science Foundation through the University of California, San Diego Materials Research Science and Engineering Center (DMR-2011924) and Graduate Research Fellowship Program (GRFP) DGE-1650112 (to E.L.W.). Support for RNA sequencing was provided by the YC Fung Chair in Bioengineering (to B.Ø.P.). M.F., A.T., and S.S.G. were supported by the National Institute of General Medical Sciences of the NIH under award R35GM118290. The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH. Publisher Copyright: Copyright © 2022 the Author(s).

PY - 2022/11/8

Y1 - 2022/11/8

N2 - UV radiation (UVR) has significant physiological effects on organisms living at or near the Earth's surface, yet the full suite of genes required for fitness of a photosynthetic organism in a UVR-rich environment remains unknown. This study reports a genome-wide fitness assessment of the genes that affect UVR tolerance under environmentally relevant UVR dosages in the model cyanobacterium Synechococcus elongatus PCC 7942. Our results highlight the importance of specific genes that encode proteins involved in DNA repair, glutathione synthesis, and the assembly and maintenance of photosystem II, as well as genes that encode hypothetical proteins and others without an obvious connection to canonical methods of UVR tolerance. Disruption of a gene that encodes a leucyl aminopeptidase (LAP) conferred the greatest UVR-specific decrease in fitness. Enzymatic assays demonstrated a strong pH-dependent affinity of the LAP for the dipeptide cysteinyl-glycine, suggesting an involvement in glutathione catabolism as a function of night-time cytosolic pH level. A low differential expression of the LAP gene under acute UVR exposure suggests that its relative importance would be overlooked in transcript-dependent screens. Subsequent experiments revealed a similar UVR-sensitivity phenotype in LAP knockouts of other organisms, indicating conservation of the functional role of LAPs in UVR tolerance.

AB - UV radiation (UVR) has significant physiological effects on organisms living at or near the Earth's surface, yet the full suite of genes required for fitness of a photosynthetic organism in a UVR-rich environment remains unknown. This study reports a genome-wide fitness assessment of the genes that affect UVR tolerance under environmentally relevant UVR dosages in the model cyanobacterium Synechococcus elongatus PCC 7942. Our results highlight the importance of specific genes that encode proteins involved in DNA repair, glutathione synthesis, and the assembly and maintenance of photosystem II, as well as genes that encode hypothetical proteins and others without an obvious connection to canonical methods of UVR tolerance. Disruption of a gene that encodes a leucyl aminopeptidase (LAP) conferred the greatest UVR-specific decrease in fitness. Enzymatic assays demonstrated a strong pH-dependent affinity of the LAP for the dipeptide cysteinyl-glycine, suggesting an involvement in glutathione catabolism as a function of night-time cytosolic pH level. A low differential expression of the LAP gene under acute UVR exposure suggests that its relative importance would be overlooked in transcript-dependent screens. Subsequent experiments revealed a similar UVR-sensitivity phenotype in LAP knockouts of other organisms, indicating conservation of the functional role of LAPs in UVR tolerance.

KW - RB-TnSeq

KW - UV radiation

KW - cyanobacteria

KW - fitness

KW - leucyl aminopeptidase

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DO - 10.1073/pnas.2211789119

M3 - Article

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SN - 0027-8424

VL - 119

SP - e2211789119

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

IS - 45

M1 - e2211789119

ER -

An unexpected role for leucyl aminopeptidase in UV tolerance revealed by a genome-wide fitness assessment in a model cyanobacterium

Abstract

Bibliographical note

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