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Nayun Kim - One of the best experts on this subject based on the ideXlab platform.
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Yeast Nucleolin Nsr1 Impedes Replication and Elevates Genome Instability at an Actively Transcribed Guanine-Rich G4 DNA-Forming Sequence.
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin (NCL) is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA-forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the cotranscriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such a G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo require the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of the RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with a guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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yeast nucleolin nsr1 impedes replication and elevates Genome Instability at an actively transcribed guanine rich g4 dna forming sequence
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA- forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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High Affinity binding of Yeast Nucleolin Nsr1 to Co-transcriptionally Formed G4 DNA Obstructs Replication and Elevates Genome Instability
2019Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:ABSTRACT A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that G4 DNA-binding is a conserved function in the yeast nucleolin Nsr1. Furthermore, we demonstrate that the Nsr1-G4 DNA complex formation results in replication obstruction and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vitro and in vivo and restores unhindered replication, overall resulting in a sharp reduction in the G4-associated Genome Instability. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and obstructing replication. AUTHOR SUMMARY Genome Instability is uniquely elevated at sequences containing multiple runs of guanines, which can fold into the unusual, four-stranded G-quadruplex (G4) DNA. In this study, we report a novel finding that a highly conserved G4 DNA binding protein Nsr1 can elevate the rate of recombination and chromosomal rearrangement occurring at a G4 DNA-forming sequence in the Genome of Saccharomyces cerevisiae. The elevated Genome Instability requires the C-terminally located RGG domain of Nsr1, which supports the high-affinity interaction between the protein and G4 DNA. The connection between G4-specific Genome Instability and the function of Nsr1 to form stable complex with G4 DNA led to the hypothesis that the high-affinity Nsr1-G4 DNA complexes can become a barrier to replication. We demonstrate here that the presence of Nsr1 in fact slows the replication past a G4 DNA-containing genomic site and that the RGG domain is required to facilitate such replication block.
Shivani Singh - One of the best experts on this subject based on the ideXlab platform.
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Yeast Nucleolin Nsr1 Impedes Replication and Elevates Genome Instability at an Actively Transcribed Guanine-Rich G4 DNA-Forming Sequence.
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin (NCL) is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA-forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the cotranscriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such a G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo require the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of the RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with a guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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yeast nucleolin nsr1 impedes replication and elevates Genome Instability at an actively transcribed guanine rich g4 dna forming sequence
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA- forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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High Affinity binding of Yeast Nucleolin Nsr1 to Co-transcriptionally Formed G4 DNA Obstructs Replication and Elevates Genome Instability
2019Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:ABSTRACT A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that G4 DNA-binding is a conserved function in the yeast nucleolin Nsr1. Furthermore, we demonstrate that the Nsr1-G4 DNA complex formation results in replication obstruction and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vitro and in vivo and restores unhindered replication, overall resulting in a sharp reduction in the G4-associated Genome Instability. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and obstructing replication. AUTHOR SUMMARY Genome Instability is uniquely elevated at sequences containing multiple runs of guanines, which can fold into the unusual, four-stranded G-quadruplex (G4) DNA. In this study, we report a novel finding that a highly conserved G4 DNA binding protein Nsr1 can elevate the rate of recombination and chromosomal rearrangement occurring at a G4 DNA-forming sequence in the Genome of Saccharomyces cerevisiae. The elevated Genome Instability requires the C-terminally located RGG domain of Nsr1, which supports the high-affinity interaction between the protein and G4 DNA. The connection between G4-specific Genome Instability and the function of Nsr1 to form stable complex with G4 DNA led to the hypothesis that the high-affinity Nsr1-G4 DNA complexes can become a barrier to replication. We demonstrate here that the presence of Nsr1 in fact slows the replication past a G4 DNA-containing genomic site and that the RGG domain is required to facilitate such replication block.
Michael Lisby - One of the best experts on this subject based on the ideXlab platform.
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topbp1 dpb11 binds dna anaphase bridges to prevent Genome Instability
Journal of Cell Biology, 2014Co-Authors: Susanne Manuela Germann, Vera Schramke, Nadine Eckertboulet, Irene Gallina, Rune Troelsgaard Pedersen, Vibe H. Oestergaard, Michael LisbyAbstract:DNA anaphase bridges are a potential source of Genome Instability that may lead to chromosome breakage or nondisjunction during mitosis. Two classes of anaphase bridges can be distinguished: DAPI-positive chromatin bridges and DAPI-negative ultrafine DNA bridges (UFBs). Here, we establish budding yeast Saccharomyces cerevisiae and the avian DT40 cell line as model systems for studying DNA anaphase bridges and show that TopBP1/Dpb11 plays an evolutionarily conserved role in their metabolism. Together with the single-stranded DNA binding protein RPA, TopBP1/Dpb11 binds to UFBs, and depletion of TopBP1/Dpb11 led to an accumulation of chromatin bridges. Importantly, the NoCut checkpoint that delays progression from anaphase to abscission in yeast was activated by both UFBs and chromatin bridges independently of Dpb11, and disruption of the NoCut checkpoint in Dpb11-depleted cells led to Genome Instability. In conclusion, we propose that TopBP1/Dpb11 prevents accumulation of anaphase bridges via stimulation of the Mec1/ATR kinase and suppression of homologous recombination.
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TopBP1/Dpb11 binds DNA anaphase bridges to prevent Genome Instability
Journal of Cell Biology, 2013Co-Authors: Susanne Manuela Germann, Vera Schramke, Irene Gallina, Rune Troelsgaard Pedersen, Vibe H. Oestergaard, Nadine Eckert-boulet, Michael LisbyAbstract:DNA anaphase bridges are a potential source of Genome Instability that may lead to chromosome breakage or nondisjunction during mitosis. Two classes of anaphase bridges can be distinguished: DAPI-positive chromatin bridges and DAPI-negative ultrafine DNA bridges (UFBs). Here, we establish budding yeast Saccharomyces cerevisiae and the avian DT40 cell line as model systems for studying DNA anaphase bridges and show that TopBP1/Dpb11 plays an evolutionarily conserved role in their metabolism. Together with the single-stranded DNA binding protein RPA, TopBP1/Dpb11 binds to UFBs, and depletion of TopBP1/Dpb11 led to an accumulation of chromatin bridges. Importantly, the NoCut checkpoint that delays progression from anaphase to abscission in yeast was activated by both UFBs and chromatin bridges independently of Dpb11, and disruption of the NoCut checkpoint in Dpb11-depleted cells led to Genome Instability. In conclusion, we propose that TopBP1/Dpb11 prevents accumulation of anaphase bridges via stimulation of the Mec1/ATR kinase and suppression of homologous recombination.
Minseon Kim - One of the best experts on this subject based on the ideXlab platform.
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Yeast Nucleolin Nsr1 Impedes Replication and Elevates Genome Instability at an Actively Transcribed Guanine-Rich G4 DNA-Forming Sequence.
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin (NCL) is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA-forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the cotranscriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such a G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo require the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of the RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with a guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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yeast nucleolin nsr1 impedes replication and elevates Genome Instability at an actively transcribed guanine rich g4 dna forming sequence
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA- forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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High Affinity binding of Yeast Nucleolin Nsr1 to Co-transcriptionally Formed G4 DNA Obstructs Replication and Elevates Genome Instability
2019Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:ABSTRACT A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that G4 DNA-binding is a conserved function in the yeast nucleolin Nsr1. Furthermore, we demonstrate that the Nsr1-G4 DNA complex formation results in replication obstruction and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vitro and in vivo and restores unhindered replication, overall resulting in a sharp reduction in the G4-associated Genome Instability. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and obstructing replication. AUTHOR SUMMARY Genome Instability is uniquely elevated at sequences containing multiple runs of guanines, which can fold into the unusual, four-stranded G-quadruplex (G4) DNA. In this study, we report a novel finding that a highly conserved G4 DNA binding protein Nsr1 can elevate the rate of recombination and chromosomal rearrangement occurring at a G4 DNA-forming sequence in the Genome of Saccharomyces cerevisiae. The elevated Genome Instability requires the C-terminally located RGG domain of Nsr1, which supports the high-affinity interaction between the protein and G4 DNA. The connection between G4-specific Genome Instability and the function of Nsr1 to form stable complex with G4 DNA led to the hypothesis that the high-affinity Nsr1-G4 DNA complexes can become a barrier to replication. We demonstrate here that the presence of Nsr1 in fact slows the replication past a G4 DNA-containing genomic site and that the RGG domain is required to facilitate such replication block.
Alexandra Berroyer - One of the best experts on this subject based on the ideXlab platform.
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Yeast Nucleolin Nsr1 Impedes Replication and Elevates Genome Instability at an Actively Transcribed Guanine-Rich G4 DNA-Forming Sequence.
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin (NCL) is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA-forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the cotranscriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such a G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo require the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of the RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with a guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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yeast nucleolin nsr1 impedes replication and elevates Genome Instability at an actively transcribed guanine rich g4 dna forming sequence
Genetics, 2020Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA- forming sequence in vivo and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. We also show that Nsr1 results in impeding replication past such G4 DNA-forming sequence. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the Genome Instability associated with guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and impeding replication.
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High Affinity binding of Yeast Nucleolin Nsr1 to Co-transcriptionally Formed G4 DNA Obstructs Replication and Elevates Genome Instability
2019Co-Authors: Shivani Singh, Alexandra Berroyer, Minseon Kim, Nayun KimAbstract:ABSTRACT A significant increase in Genome Instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and Genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin NCL is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the Genome Instability has not been previously investigated. We show here that G4 DNA-binding is a conserved function in the yeast nucleolin Nsr1. Furthermore, we demonstrate that the Nsr1-G4 DNA complex formation results in replication obstruction and is a major factor in inducing the Genome Instability associated with the co-transcriptionally formed G4 DNA in the yeast Genome. The G4-associated Genome Instability and the G4 DNA-binding in vivo requires the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vitro and in vivo and restores unhindered replication, overall resulting in a sharp reduction in the G4-associated Genome Instability. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs Genome stability by precluding the access of G4-resolving proteins and obstructing replication. AUTHOR SUMMARY Genome Instability is uniquely elevated at sequences containing multiple runs of guanines, which can fold into the unusual, four-stranded G-quadruplex (G4) DNA. In this study, we report a novel finding that a highly conserved G4 DNA binding protein Nsr1 can elevate the rate of recombination and chromosomal rearrangement occurring at a G4 DNA-forming sequence in the Genome of Saccharomyces cerevisiae. The elevated Genome Instability requires the C-terminally located RGG domain of Nsr1, which supports the high-affinity interaction between the protein and G4 DNA. The connection between G4-specific Genome Instability and the function of Nsr1 to form stable complex with G4 DNA led to the hypothesis that the high-affinity Nsr1-G4 DNA complexes can become a barrier to replication. We demonstrate here that the presence of Nsr1 in fact slows the replication past a G4 DNA-containing genomic site and that the RGG domain is required to facilitate such replication block.