Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space

Long-term space missions will expose crew members, their cells as well as their microbiomes to prolonged periods of microgravity and ionizing radiation, environmental stressors for which almost no earth-based organisms have evolved to survive. Despite the importance of maintaining genomic integrity,...

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Autores principales:	Aaron H Rosenstein, Virginia K Walker
Formato:	article
Lenguaje:	EN
Publicado:	Frontiers Media S.A. 2021
Materias:	klenow exonuclease DNA repair microgravity base substitutions DNA deletions next generation sequencing Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/45fb33bc76d9455080930a0a5ab7f73f
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id	oai:doaj.org-article:45fb33bc76d9455080930a0a5ab7f73f
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spelling	oai:doaj.org-article:45fb33bc76d9455080930a0a5ab7f73f2021-12-01T10:35:25ZFidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space2296-634X10.3389/fcell.2021.702849https://doaj.org/article/45fb33bc76d9455080930a0a5ab7f73f2021-11-01T00:00:00Zhttps://www.frontiersin.org/articles/10.3389/fcell.2021.702849/fullhttps://doaj.org/toc/2296-634XLong-term space missions will expose crew members, their cells as well as their microbiomes to prolonged periods of microgravity and ionizing radiation, environmental stressors for which almost no earth-based organisms have evolved to survive. Despite the importance of maintaining genomic integrity, the impact of these stresses on DNA polymerase-mediated replication and repair has not been fully explored. DNA polymerase fidelity and replication rates were assayed under conditions of microgravity generated by parabolic flight and compared to earth-like gravity. Upon commencement of a parabolic arc, primed synthetic single-stranded DNA was used as a template for one of two enzymes (Klenow fragment exonuclease+/−; with and without proofreading exonuclease activity, respectively) and were quenched immediately following the 20 s microgravitational period. DNA polymerase error rates were determined with an algorithm developed to identify experimental mutations. In microgravity Klenow exonuclease+ showed a median 1.1-fold per-base decrease in polymerization fidelity for base substitutions when compared to earth-like gravity (p = 0.02), but in the absence of proofreading activity, a 2.4-fold decrease was observed (p = 1.98 × 10−11). Similarly, 1.1-fold and 1.5-fold increases in deletion frequencies in the presence or absence of exonuclease activity (p = 1.51 × 10−7 and p = 8.74 × 10−13), respectively, were observed in microgravity compared to controls. The development of this flexible semi-autonomous payload system coupled with genetic and bioinformatic approaches serves as a proof-of-concept for future space health research.Aaron H RosensteinVirginia K WalkerFrontiers Media S.A.articleklenow exonucleaseDNA repairmicrogravitybase substitutionsDNA deletionsnext generation sequencingBiology (General)QH301-705.5ENFrontiers in Cell and Developmental Biology, Vol 9 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	klenow exonuclease DNA repair microgravity base substitutions DNA deletions next generation sequencing Biology (General) QH301-705.5
spellingShingle	klenow exonuclease DNA repair microgravity base substitutions DNA deletions next generation sequencing Biology (General) QH301-705.5 Aaron H Rosenstein Virginia K Walker Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space
description	Long-term space missions will expose crew members, their cells as well as their microbiomes to prolonged periods of microgravity and ionizing radiation, environmental stressors for which almost no earth-based organisms have evolved to survive. Despite the importance of maintaining genomic integrity, the impact of these stresses on DNA polymerase-mediated replication and repair has not been fully explored. DNA polymerase fidelity and replication rates were assayed under conditions of microgravity generated by parabolic flight and compared to earth-like gravity. Upon commencement of a parabolic arc, primed synthetic single-stranded DNA was used as a template for one of two enzymes (Klenow fragment exonuclease+/−; with and without proofreading exonuclease activity, respectively) and were quenched immediately following the 20 s microgravitational period. DNA polymerase error rates were determined with an algorithm developed to identify experimental mutations. In microgravity Klenow exonuclease+ showed a median 1.1-fold per-base decrease in polymerization fidelity for base substitutions when compared to earth-like gravity (p = 0.02), but in the absence of proofreading activity, a 2.4-fold decrease was observed (p = 1.98 × 10−11). Similarly, 1.1-fold and 1.5-fold increases in deletion frequencies in the presence or absence of exonuclease activity (p = 1.51 × 10−7 and p = 8.74 × 10−13), respectively, were observed in microgravity compared to controls. The development of this flexible semi-autonomous payload system coupled with genetic and bioinformatic approaches serves as a proof-of-concept for future space health research.
format	article
author	Aaron H Rosenstein Virginia K Walker
author_facet	Aaron H Rosenstein Virginia K Walker
author_sort	Aaron H Rosenstein
title	Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space
title_short	Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space
title_full	Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space
title_fullStr	Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space
title_full_unstemmed	Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space
title_sort	fidelity of a bacterial dna polymerase in microgravity, a model for human health in space
publisher	Frontiers Media S.A.
publishDate	2021
url	https://doaj.org/article/45fb33bc76d9455080930a0a5ab7f73f
work_keys_str_mv	AT aaronhrosenstein fidelityofabacterialdnapolymeraseinmicrogravityamodelforhumanhealthinspace AT virginiakwalker fidelityofabacterialdnapolymeraseinmicrogravityamodelforhumanhealthinspace
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Fidelity of a Bacterial DNA Polymerase in Microgravity, a Model for Human Health in Space

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