Additional file 3. Mutations in ORFs of evolved non-mutator strains, grouped according to their p... more Additional file 3. Mutations in ORFs of evolved non-mutator strains, grouped according to their predicted effect.
Additional file 12. Correlation of point mutations located to ORF, and putative promoter and term... more Additional file 12. Correlation of point mutations located to ORF, and putative promoter and terminator regions and transcriptomic changes in evolved yeast strains.
One sentence summary: The stability of budding and fission yeast genomes is influenced by two con... more One sentence summary: The stability of budding and fission yeast genomes is influenced by two contradictory factors: (1) the need to be fully functional, which is ensured through the replication fidelity pathways of nuclear and mitochondrial genomes through sensing and repairing DNA damage, through precise chromosome segregation during cell division; and (2) the need to acquire changes for adaptation to environmental challenges. Editor: Antoine Danchin Cells are constantly confronted with endogenous and exogenous factors that affect their genomes. Eons of evolution have allowed the cellular mechanisms responsible for preserving the genome to adjust for achieving contradictory objectives: to maintain the genome unchanged and to acquire mutations that allow adaptation to environmental changes. One evolutionary mechanism that has been refined for survival is genetic variation. In this review, we describe the mechanisms responsible for two biological processes: genome maintenance and mu...
One sentence summary: The stability of budding and fission yeast genomes is influenced by two con... more One sentence summary: The stability of budding and fission yeast genomes is influenced by two contradictory factors: (1) the need to be fully functional, which is ensured through the replication fidelity pathways of nuclear and mitochondrial genomes through sensing and repairing DNA damage, through precise chromosome segregation during cell division; and (2) the need to acquire changes for adaptation to environmental challenges. Editor: Antoine Danchin Cells are constantly confronted with endogenous and exogenous factors that affect their genomes. Eons of evolution have allowed the cellular mechanisms responsible for preserving the genome to adjust for achieving contradictory objectives: to maintain the genome unchanged and to acquire mutations that allow adaptation to environmental changes. One evolutionary mechanism that has been refined for survival is genetic variation. In this review, we describe the mechanisms responsible for two biological processes: genome maintenance and mu...
The yeast Saccharomyces cerevisiae is a unicellular organism commonly used as a model to explain ... more The yeast Saccharomyces cerevisiae is a unicellular organism commonly used as a model to explain mechanisms of aging in multicellular organisms. It is used as a model organism for both replicative and chronological aging. Replicative aging is defined as the number of daughter cells produced by an individual cell during its life. A widely accepted hypothesis assumes that replicative aging of yeast is related to the existence of a so called "senescence factor" that gradually accumulates in the mother cell, which consequently leads to its death. One of the earliest proposed "senescence factors" were extrachromosomal rDNA circles (ERCs). However, their role in the regulation of the replicative lifespan is somewhat controversial and subject to discussion. In this paper, we propose a more comprehensive approach to this problem by analysing the length of life and the correlation between the cell size and the replicative lifespan of yeast cells with different level of ERCs, i.e. Δrad52 and Δsgs1 mutants. This analysis shows that it is not the accumulation of ERCs but genomic instability and hypertrophy that play an important role in the regulation of reproductive potential and total lifespan of the S. cerevisiae yeast. However, these two factors have a different impact on various phases of the yeast cell life, i.e. reproductive and post-reproductive phases.
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, 2006
Besides its role as a major recycler of unfolded or otherwise damaged intracellular proteins, the... more Besides its role as a major recycler of unfolded or otherwise damaged intracellular proteins, the 26S proteasome functions as a regulator of many vital cellular processes and is postulated as a target for antitumor drugs. It has previously been shown that dysfunction of the catalytic core of the 26S proteasome, the 20S proteasome, causes a moderate increase in the frequency of spontaneous mutations in yeast [A. Podlaska, J. McIntyre, A. Skoneczna, E. Sledziewska-Gojska, The link between proteasome activity and postreplication DNA repair in Saccharomyces cerevisiae. Mol. Microbiol. 49 (2003) 1321-1332]. Here we show the results of genetic analysis, which indicate that the mutator phenotype caused by the deletion of UMP1, encoding maturase of 20S proteasome, involves members of the RAD6 epistasis group. The great majority of mutations occurring spontaneously in yeast cells deficient in 20S proteasome function are connected with the unique Rad6/Rad18-dependent error-prone translesion DNA synthesis (TLS) requiring the activities of both TLS polymerases: Pol eta and Pol zeta. Our results suggest the involvement of proteasomal activity in the limitation of this unique error-prone TLS mechanism in wild-type cells. On the other hand, we found that the mutator phenotypes caused by deficiency in Rad18 and Rad6, are largely alleviated by defects in proteasome activities. Since the mutator phenotypes produced by deletion of RAD6 and RAD18 require Pol zeta and Siz1/Ubc9-dependent sumoylation of PCNA, our results suggest that proteasomal dysfunction limits sumoylation-dependent error-prone activity of Pol zeta. Taken together, our findings strongly support the idea that proteolytic activity is involved in modulating the balance between TLS mechanisms functioning during DNA replication in S. cerevisiae.
The integrity of a cellular genome may be endangered in many ways. However, cells are not powerle... more The integrity of a cellular genome may be endangered in many ways. However, cells are not powerless and indifferent to their future fate; they are prepared to solve various problems concerning their genome to enable their survival. Maintaining a stable genome is a complicated task, even without any external problems such as genotoxic agents, irradiation, or any other harmful treatment to which cells are exposed in their environment. Inaccuracies in replication alone may lead to changes in DNA sequence, i.e., to mutations. When cells are exposed to stressful conditions that cause DNA lesions, the frequency of mutations increases. Moreover, unrepaired DNA damage may lead to the replication fork block that delays the replication round until the damage is repaired. However, if for some reason the DNA damage in the replication fork remains unrepaired, it may lead to DNA breakage causing DNA rearrangements or to fork collapse followed by cell death. Consequently, DNA damage is dangerous for replicating cells. What happens when DNA damage appears after a replication round has been successfully finished? Various DNA lesions may disturb the transcription efficiency of the affected genes. DNA strand breaks may generate problems during chromosome segregation because such lesions may lead to an unequal DNA distribution to daughter cells.
We have shown previously that deletion of the Saccharomyces cerevisiae UMP1 gene encoding the 20S... more We have shown previously that deletion of the Saccharomyces cerevisiae UMP1 gene encoding the 20S proteasome maturase causes sensitivity to UV radiation. In the current report, we have extended this finding to show that mutations specifically compromising chymotrypsin-like or trypsin-like activity of 20S proteasome peptidases also result in increased UV sensitivity. We have also established that mutations affecting proteasome activity, namely ump1Delta, pre2-K108R and pup1-T20A, result in spontaneous and UV-induced mutator phenotypes. To elucidate the origin of these DNA repair phenotypes of the proteasomal mutants, we performed epistasis analysis, with respect to UV sensitivity, using yeast strains with the UMP1 deletion in different DNA repair backgrounds. We show that UMP1 is not epistatic to RAD23 and RAD2, which are involved in the nucleotide excision repair (NER) pathway. Instead, our results indicate that UMP1 as well as PUP1 and PRE2 (encoding catalytic subunits of 20S proteasome) belong to an epistatic group of genes functioning in post-replication DNA repair (PRR) and are hypostatic to RAD18, which, in complex with RAD6, plays a central role in PRR. We also show that UMP1 is epistatic to REV3 and RAD30, although the relationship of UMP1 with these genes is different.
Biochimica et biophysica acta. Molecular cell research, Jun 1, 2023
The DNA double-strand breaks are particularly deleterious, especially when an error-free repair p... more The DNA double-strand breaks are particularly deleterious, especially when an error-free repair pathway is unavailable, enforcing the error-prone recombination pathways to repair the lesion. Cells can resume the cell cycle but at the expense of decreased viability due to genome rearrangements. One of the major players involved in recombinational repair of DNA damage is Rad51 recombinase, a protein responsible for presynaptic complex formation. We previously showed that an increased level of this protein promotes the usage of illegitimate recombination. Here we show that the level of Rad51 is regulated via the ubiquitin-dependent proteolytic pathway. The ubiquitination of Rad51 depends on multiple E3 enzymes, including SUMO-targeted ubiquitin ligases. We also demonstrate that Rad51 can be modified by both ubiquitin and SUMO. Moreover, its modification with ubiquitin may lead to opposite effects: degradation dependent on Rad6, Rad18, Slx8, Dia2, and the anaphase-promoting complex, or stabilization dependent on Rsp5. We also show that post-translational modifications with SUMO and ubiquitin affect Rad51's ability to form and disassemble DNA repair foci, respectively, influencing cell cycle progression and cell viability in genotoxic stress conditions. Our data suggest the existence of a complex E3 ligases network that regulates Rad51 recombinase's turnover, its molecular activity, and access to DNA, limiting it to the proportions optimal for the actual cell cycle stage and growth conditions, e.g., stress. Dysregulation of this network would result in a drop in cell viability due to uncontrolled genome rearrangement in the yeast cells. In mammals would promote the development of genetic diseases and cancer.
Additional file 3. Mutations in ORFs of evolved non-mutator strains, grouped according to their p... more Additional file 3. Mutations in ORFs of evolved non-mutator strains, grouped according to their predicted effect.
Additional file 12. Correlation of point mutations located to ORF, and putative promoter and term... more Additional file 12. Correlation of point mutations located to ORF, and putative promoter and terminator regions and transcriptomic changes in evolved yeast strains.
One sentence summary: The stability of budding and fission yeast genomes is influenced by two con... more One sentence summary: The stability of budding and fission yeast genomes is influenced by two contradictory factors: (1) the need to be fully functional, which is ensured through the replication fidelity pathways of nuclear and mitochondrial genomes through sensing and repairing DNA damage, through precise chromosome segregation during cell division; and (2) the need to acquire changes for adaptation to environmental challenges. Editor: Antoine Danchin Cells are constantly confronted with endogenous and exogenous factors that affect their genomes. Eons of evolution have allowed the cellular mechanisms responsible for preserving the genome to adjust for achieving contradictory objectives: to maintain the genome unchanged and to acquire mutations that allow adaptation to environmental changes. One evolutionary mechanism that has been refined for survival is genetic variation. In this review, we describe the mechanisms responsible for two biological processes: genome maintenance and mu...
One sentence summary: The stability of budding and fission yeast genomes is influenced by two con... more One sentence summary: The stability of budding and fission yeast genomes is influenced by two contradictory factors: (1) the need to be fully functional, which is ensured through the replication fidelity pathways of nuclear and mitochondrial genomes through sensing and repairing DNA damage, through precise chromosome segregation during cell division; and (2) the need to acquire changes for adaptation to environmental challenges. Editor: Antoine Danchin Cells are constantly confronted with endogenous and exogenous factors that affect their genomes. Eons of evolution have allowed the cellular mechanisms responsible for preserving the genome to adjust for achieving contradictory objectives: to maintain the genome unchanged and to acquire mutations that allow adaptation to environmental changes. One evolutionary mechanism that has been refined for survival is genetic variation. In this review, we describe the mechanisms responsible for two biological processes: genome maintenance and mu...
The yeast Saccharomyces cerevisiae is a unicellular organism commonly used as a model to explain ... more The yeast Saccharomyces cerevisiae is a unicellular organism commonly used as a model to explain mechanisms of aging in multicellular organisms. It is used as a model organism for both replicative and chronological aging. Replicative aging is defined as the number of daughter cells produced by an individual cell during its life. A widely accepted hypothesis assumes that replicative aging of yeast is related to the existence of a so called "senescence factor" that gradually accumulates in the mother cell, which consequently leads to its death. One of the earliest proposed "senescence factors" were extrachromosomal rDNA circles (ERCs). However, their role in the regulation of the replicative lifespan is somewhat controversial and subject to discussion. In this paper, we propose a more comprehensive approach to this problem by analysing the length of life and the correlation between the cell size and the replicative lifespan of yeast cells with different level of ERCs, i.e. Δrad52 and Δsgs1 mutants. This analysis shows that it is not the accumulation of ERCs but genomic instability and hypertrophy that play an important role in the regulation of reproductive potential and total lifespan of the S. cerevisiae yeast. However, these two factors have a different impact on various phases of the yeast cell life, i.e. reproductive and post-reproductive phases.
Mutation Research: Fundamental And Molecular Mechanisms Of Mutagenesis, 2006
Besides its role as a major recycler of unfolded or otherwise damaged intracellular proteins, the... more Besides its role as a major recycler of unfolded or otherwise damaged intracellular proteins, the 26S proteasome functions as a regulator of many vital cellular processes and is postulated as a target for antitumor drugs. It has previously been shown that dysfunction of the catalytic core of the 26S proteasome, the 20S proteasome, causes a moderate increase in the frequency of spontaneous mutations in yeast [A. Podlaska, J. McIntyre, A. Skoneczna, E. Sledziewska-Gojska, The link between proteasome activity and postreplication DNA repair in Saccharomyces cerevisiae. Mol. Microbiol. 49 (2003) 1321-1332]. Here we show the results of genetic analysis, which indicate that the mutator phenotype caused by the deletion of UMP1, encoding maturase of 20S proteasome, involves members of the RAD6 epistasis group. The great majority of mutations occurring spontaneously in yeast cells deficient in 20S proteasome function are connected with the unique Rad6/Rad18-dependent error-prone translesion DNA synthesis (TLS) requiring the activities of both TLS polymerases: Pol eta and Pol zeta. Our results suggest the involvement of proteasomal activity in the limitation of this unique error-prone TLS mechanism in wild-type cells. On the other hand, we found that the mutator phenotypes caused by deficiency in Rad18 and Rad6, are largely alleviated by defects in proteasome activities. Since the mutator phenotypes produced by deletion of RAD6 and RAD18 require Pol zeta and Siz1/Ubc9-dependent sumoylation of PCNA, our results suggest that proteasomal dysfunction limits sumoylation-dependent error-prone activity of Pol zeta. Taken together, our findings strongly support the idea that proteolytic activity is involved in modulating the balance between TLS mechanisms functioning during DNA replication in S. cerevisiae.
The integrity of a cellular genome may be endangered in many ways. However, cells are not powerle... more The integrity of a cellular genome may be endangered in many ways. However, cells are not powerless and indifferent to their future fate; they are prepared to solve various problems concerning their genome to enable their survival. Maintaining a stable genome is a complicated task, even without any external problems such as genotoxic agents, irradiation, or any other harmful treatment to which cells are exposed in their environment. Inaccuracies in replication alone may lead to changes in DNA sequence, i.e., to mutations. When cells are exposed to stressful conditions that cause DNA lesions, the frequency of mutations increases. Moreover, unrepaired DNA damage may lead to the replication fork block that delays the replication round until the damage is repaired. However, if for some reason the DNA damage in the replication fork remains unrepaired, it may lead to DNA breakage causing DNA rearrangements or to fork collapse followed by cell death. Consequently, DNA damage is dangerous for replicating cells. What happens when DNA damage appears after a replication round has been successfully finished? Various DNA lesions may disturb the transcription efficiency of the affected genes. DNA strand breaks may generate problems during chromosome segregation because such lesions may lead to an unequal DNA distribution to daughter cells.
We have shown previously that deletion of the Saccharomyces cerevisiae UMP1 gene encoding the 20S... more We have shown previously that deletion of the Saccharomyces cerevisiae UMP1 gene encoding the 20S proteasome maturase causes sensitivity to UV radiation. In the current report, we have extended this finding to show that mutations specifically compromising chymotrypsin-like or trypsin-like activity of 20S proteasome peptidases also result in increased UV sensitivity. We have also established that mutations affecting proteasome activity, namely ump1Delta, pre2-K108R and pup1-T20A, result in spontaneous and UV-induced mutator phenotypes. To elucidate the origin of these DNA repair phenotypes of the proteasomal mutants, we performed epistasis analysis, with respect to UV sensitivity, using yeast strains with the UMP1 deletion in different DNA repair backgrounds. We show that UMP1 is not epistatic to RAD23 and RAD2, which are involved in the nucleotide excision repair (NER) pathway. Instead, our results indicate that UMP1 as well as PUP1 and PRE2 (encoding catalytic subunits of 20S proteasome) belong to an epistatic group of genes functioning in post-replication DNA repair (PRR) and are hypostatic to RAD18, which, in complex with RAD6, plays a central role in PRR. We also show that UMP1 is epistatic to REV3 and RAD30, although the relationship of UMP1 with these genes is different.
Biochimica et biophysica acta. Molecular cell research, Jun 1, 2023
The DNA double-strand breaks are particularly deleterious, especially when an error-free repair p... more The DNA double-strand breaks are particularly deleterious, especially when an error-free repair pathway is unavailable, enforcing the error-prone recombination pathways to repair the lesion. Cells can resume the cell cycle but at the expense of decreased viability due to genome rearrangements. One of the major players involved in recombinational repair of DNA damage is Rad51 recombinase, a protein responsible for presynaptic complex formation. We previously showed that an increased level of this protein promotes the usage of illegitimate recombination. Here we show that the level of Rad51 is regulated via the ubiquitin-dependent proteolytic pathway. The ubiquitination of Rad51 depends on multiple E3 enzymes, including SUMO-targeted ubiquitin ligases. We also demonstrate that Rad51 can be modified by both ubiquitin and SUMO. Moreover, its modification with ubiquitin may lead to opposite effects: degradation dependent on Rad6, Rad18, Slx8, Dia2, and the anaphase-promoting complex, or stabilization dependent on Rsp5. We also show that post-translational modifications with SUMO and ubiquitin affect Rad51's ability to form and disassemble DNA repair foci, respectively, influencing cell cycle progression and cell viability in genotoxic stress conditions. Our data suggest the existence of a complex E3 ligases network that regulates Rad51 recombinase's turnover, its molecular activity, and access to DNA, limiting it to the proportions optimal for the actual cell cycle stage and growth conditions, e.g., stress. Dysregulation of this network would result in a drop in cell viability due to uncontrolled genome rearrangement in the yeast cells. In mammals would promote the development of genetic diseases and cancer.
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Papers by Adrianna Skoneczna