Gene dosage compensation of rRNA transcript levels in Arabidopsis thaliana lines with reduced ribosomal gene copy number

The Plant Cell ◽

10.1093/plcell/koab020 ◽

2021 ◽

Author(s):

Francesca B Lopez ◽

Antoine Fort ◽

Luca Tadini ◽

Aline V Probst ◽

Marcus McHale ◽

...

Keyword(s):

Arabidopsis Thaliana ◽

Genome Editing ◽

Dosage Compensation ◽

Copy Number ◽

Gene Dosage ◽

Rrna Genes ◽

Gene Copy ◽

Transcript Levels ◽

Rdna Copy ◽

Rdna Copy Number

Abstract The 45S rRNA genes (rDNA) are amongst the largest repetitive elements in eukaryotic genomes. rDNA consists of tandem arrays of rRNA genes, many of which are transcriptionally silenced. Silent rDNA repeats may act as ‘back-up’ copies for ribosome biogenesis and have nuclear organization roles. Through Cas9-mediated genome editing in the Arabidopsis thaliana female gametophyte we reduced 45S rDNA copy number to a plateau of ∼10%. Two independent lines had rDNA copy numbers reduced by up to 90% at the T7 generation, named Low Copy Number (LCN) lines. Despite drastic reduction of rDNA copies, rRNA transcriptional rates and steady-state levels remained the same as wild type plants. Gene dosage compensation of rRNA transcript levels was associated with reduction of silencing histone marks at rDNA loci and altered Nucleolar Organiser Region 2 organization. While overall genome integrity of LCN lines appears unaffected, a chromosome segmental duplication occurred in one of the lines. Transcriptome analysis of LCN seedlings identified several shared dysregulated genes and pathways in both independent lines. Cas9 genome editing of rRNA repeats to generate LCN lines provides a powerful technique to elucidate rDNA dosage compensation mechanisms and impacts of low rDNA copy number on genome stability, development, and cellular processes.

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Unstable inheritance of 45S rRNA genes in Arabidopsis thaliana

10.1101/088088 ◽

2016 ◽

Author(s):

Fernando A. Rabanal ◽

Viktoria Nizhynska ◽

Terezie Mandáková ◽

Polina Yu. Novikova ◽

Martin A. Lysak ◽

...

Keyword(s):

Arabidopsis Thaliana ◽

Copy Number ◽

Association Studies ◽

Recombinant Inbred Lines ◽

Gene Copy Number ◽

Rrna Genes ◽

Gene Copy ◽

Nucleolar Organizer Regions ◽

Rrna Gene ◽

Genome Wide Association Studies

AbstractThe considerable genome size variation in Arabidopsis thaliana has been shown largely to be due to copy number variation (CNV) in 45S ribosomal RNA (rRNA) genes. Surprisingly, attempts to map this variation by means of genome-wide association studies (GWAS) failed to identify either of the two likely sources, namely the nucleolar organizer regions (NORs). Instead, GWAS implicated a trans-acting locus, as if rRNA CNV was a phenotype rather than a genotype. To explain these results, we investigated the inheritance and stability of rRNA gene copy number using the variety of genetic resources available in A. thaliana — F2 crosses, recombinant inbred lines, the multiparent advanced generation inter-cross population, and mutation accumulation lines. Our results clearly show that rRNA gene CNV can be mapped to the NORs themselves, with both loci contributing equally to the variation. However, NOR size is unstably inherited, and dramatic copy number changes are visible already within tens of generations, which explains why it is not possible to map the NORs using GWAS. We did not find any evidence of trans-acting loci in crosses, which is also expected since changes due to such loci would take very many generations to manifest themselves. rRNA gene copy number is thus an interesting example of “missing heritability” — a trait that is heritable in pedigrees, but not in the general population.

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Копийность рибосомной ДНК (рДНК) в геномах женщин как фактор успешности ЭКО и наличия осложнений беременности

Nauchno-prakticheskii zhurnal «Medicinskaia genetika» ◽

10.25557/2073-7998.2019.11.14-25 ◽

2019 ◽

pp. 14-25

Author(s):

Л.Н. Пороховник ◽

Н.Н. Вейко ◽

Е.С. Ершова ◽

А.А. Полеткина ◽

Г.В. Шмарина ◽

...

Keyword(s):

Cell Death ◽

Stress Resistance ◽

Copy Number ◽

Gene Copy ◽

Death Process ◽

Copy Numbers ◽

The Individual ◽

Complicated Pregnancy ◽

Rdna Copy ◽

Rdna Copy Number

Определено количество копий рибосомных повторов (рДНК) в геномах женщин с нормальной и осложненной беременностью, а также женщин, подвергшихся процедуре экстракорпорального оплодотворения (ЭКО). Кроме того, измеряли содержание GC-богатой рДНК в образцах внеклеточной ДНК (вкДНК), полученных от женщин с нормальной и осложненной беременностью. Показано, что геномы более половины женщин с патологией беременности содержали либо больше, либо меньше копий рДНК, чем у любой женщины из контрольной группы. Также обнаружено более высокое содержание рДНК во вкДНК пациенток с осложненной беременностью, что свидетельствует о наличии хронического процесса аномальной гибели клеток в группе женщин с патологией беременности. Можно сделать принципиальный вывод: поскольку беременность является тяжелой нагрузкой на организм женщины, для успешного вынашивания требуется сбалансированный биогенез рибосом. Женщины как с низкой, так и с очень высокой копийностью рДНК имеют более высокую вероятность повышенного уровня апоптоза и попадания в группу риска. Параметр «число копий рДНК в геноме женщины» может служить дополнительным прогностическим маркером потенциальных осложнений беременности у женщины. Женщины с низким или высоким количеством копий рибосомных генов в геноме нуждаются в более внимательном ведении беременности. Показатели количества копий рДНК в геномах женщин с неудачными попытками ЭКО были значимо ниже, чем в геномах двух остальных групп. Этот факт говорит о том, что копийность рДНК в геноме является одним из факторов, влияющих на успех процедуры ЭКО. Если индвивидуальное число копий рДНК в геноме женщины меньше, чем 330, высок риск неудачного ЭКО. Необходимы дальнейшие исследования данного вопроса. As pregnancy is a stressful load for a woman, any stress-resistance factor is relevant to it. According to recent reports, ribosomal gene copy number in the genome is associated with the individual stress-resistance. We determined copy numbers of ribosomal DNA (rDNA) in genomes of pregnant women with normal and complicated pregnancy, and women after in vitro fertilization (IVF) procedure. We also measured the contents of GC-rich rDNA in cell-free DNA (cfDNA) derived from normal controls and complicated pregnancy cases. We have shown that genomes of more than a half of DNA samples from women with pregnancy pathology harbor either more, or less rDNA copies than any woman from the control group. We also found higher rDNA contents in cfDNA isolated from complicated pregnancy cases suggesting the presence of a permanent cell death process in pathology cases. A principal conclusion can be made: women with low rDNA copy numbers and with very high numbers can have higher cell death rates and belong to the risk group. The parameter «rDNA copy number in woman’s genome» can be an additional prognostic marker for eventual pregnancy complications in the woman. The numbers of rDNA copies in the genomes of women with failed IVF attempts was significantly lower than in the genomes of patients with succesfull outcome, suggesting that rDNA copy number in the genome is one of the factors that affect the success of the IVF procedure. If the individual rDNA copy number is under 330, the risk of IVF failure is high. Further studies are warranted.

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Transgenerational dynamics of rDNA copy number in Drosophila male germline stem cells

10.1101/199679 ◽

2017 ◽

Author(s):

Kevin L. Lu ◽

Jonathan O. Nelson ◽

Natalie Warsinger-Pepe ◽

Yukiko M. Yamashita

Keyword(s):

Copy Number ◽

Replicative Senescence ◽

Yeast Cells ◽

Rrna Genes ◽

Germline Stem Cell ◽

Genetic Elements ◽

Male Germline ◽

Rdna Loci ◽

Rdna Copy ◽

Rdna Copy Number

AbstractrDNA loci, composed of hundreds of tandemly duplicated arrays of rRNA genes, are known to be among the most unstable genetic elements due to their repetitive nature. rDNA instability underlies aging (replicative senescence) in yeast cells, however, its contribution to the aging of multicellular organisms is poorly understood. In this study, we investigate the dynamics of rDNA loci during aging in the Drosophila male germline stem cell (GSC) lineage, and show that rDNA copy number decreases during aging. Our study further reveals that this age-dependent decrease in rDNA copy number is heritable from generation to generation, yet GSCs in animals that inherit reduced rDNA copy number are capable of recovering normal rDNA copy number. Based on these findings, we propose that rDNA loci are dynamic genetic elements, where rDNA copy number changes dynamically yet is maintained through a recovery mechanism in the germline.

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The correlation between rDNA copy number and genome size in eukaryotes

Genome ◽

10.1139/g02-103 ◽

2003 ◽

Vol 46 (1) ◽

pp. 48-50 ◽

Cited By ~ 270

Author(s):

Cheryl D Prokopowich ◽

T Ryan Gregory ◽

Teresa J Crease

Keyword(s):

Genome Size ◽

Copy Number ◽

Positive Relationship ◽

Convincing Evidence ◽

Rrna Genes ◽

Strong Positive Relationship ◽

C Value ◽

Two Parameters ◽

Rdna Copy ◽

Rdna Copy Number

Both rDNA gene multiplicity and genome size vary widely among eukaryotes. For some time, there has been debate regarding any possible relationship between these two parameters. The present study uses data on genome size and rDNA copy number for 162 species of plants and animals to test the association between genome size and rDNA copy number, and provides the first convincing evidence of a strong positive relationship between the two within and among these two groups of organisms. No simple explanations exist for this relationship, but it is nevertheless of clear relevance from both practical and theoretical perspectives.Key words: rRNA genes, C value, genome size.

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Transgenerational dynamics of rDNA copy number in Drosophila male germline stem cells

eLife ◽

10.7554/elife.32421 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 20

Author(s):

Kevin L Lu ◽

Jonathan O Nelson ◽

George J Watase ◽

Natalie Warsinger-Pepe ◽

Yukiko M Yamashita

Keyword(s):

Copy Number ◽

Replicative Senescence ◽

Yeast Cells ◽

Rrna Genes ◽

Germline Stem Cell ◽

Genetic Elements ◽

Male Germline ◽

Rdna Loci ◽

Rdna Copy ◽

Rdna Copy Number

rDNA loci, composed of hundreds of tandemly duplicated arrays of rRNA genes, are known to be among the most unstable genetic elements due to their repetitive nature. rDNA instability underlies aging (replicative senescence) in yeast cells, however, its contribution to the aging of multicellular organisms is poorly understood. In this study, we investigate the dynamics of rDNA loci during aging in the Drosophila male germline stem cell (GSC) lineage, and show that rDNA copy number decreases during aging. Our study further reveals that this age-dependent decrease in rDNA copy number is heritable from generation to generation, yet GSCs in young animals that inherited reduced rDNA copy number are capable of recovering normal rDNA copy number. Based on these findings, we propose that rDNA loci are dynamic genetic elements, where rDNA copy number changes dynamically yet is maintained through a recovery mechanism in the germline.

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Transcription of ribosomal genes can cause nondisjunction

The Journal of Cell Biology ◽

10.1083/jcb.200511129 ◽

2006 ◽

Vol 173 (6) ◽

pp. 893-903 ◽

Cited By ~ 26

Author(s):

Felix Machín ◽

Jordi Torres-Rosell ◽

Giacomo De Piccoli ◽

Jesús A. Carballo ◽

Rita S. Cha ◽

...

Keyword(s):

Ribosomal Rna ◽

Copy Number ◽

Ribosomal Genes ◽

Rrna Genes ◽

Normal Length ◽

Very High ◽

Rdna Copy ◽

Rdna Copy Number ◽

Genetic Rearrangements ◽

Chromosome Nondisjunction

Mitotic disjunction of the repetitive ribosomal DNA (rDNA) involves specialized segregation mechanisms dependent on the conserved phosphatase Cdc14. The reason behind this requirement is unknown. We show that rDNA segregation requires Cdc14 partly because of its physical length but most importantly because a fraction of ribosomal RNA (rRNA) genes are transcribed at very high rates. We show that cells cannot segregate rDNA without Cdc14 unless they undergo genetic rearrangements that reduce rDNA copy number. We then demonstrate that cells with normal length rDNA arrays can segregate rDNA in the absence of Cdc14 as long as rRNA genes are not transcribed. In addition, our study uncovers an unexpected role for the replication barrier protein Fob1 in rDNA segregation that is independent of Cdc14. These findings demonstrate that highly transcribed loci can cause chromosome nondisjunction.

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Faculty Opinions recommendation of rDNA Copy Number Variants Are Frequent Passenger Mutations in Saccharomyces cerevisiae Deletion Collections and de Novo Transformants.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.726580404.793522980 ◽

2016 ◽

Author(s):

Judith Berman

Keyword(s):

Saccharomyces Cerevisiae ◽

Copy Number ◽

De Novo ◽

Copy Number Variants ◽

Passenger Mutations ◽

Rdna Copy ◽

Rdna Copy Number

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On the Mode of Gene-Dosage Compensation in Drosophila

Genetics ◽

10.1093/genetics/145.3.729 ◽

1997 ◽

Vol 145 (3) ◽

pp. 729-736

Author(s):

Irina Arkhipova ◽

Jingjing Li ◽

Matthew Meselson

Keyword(s):

X Chromosome ◽

Dosage Compensation ◽

Gene Dosage ◽

Gene Activity ◽

Gene Copy ◽

Drosophila Pseudoobscura ◽

A Site ◽

Reduced Activity ◽

Per Gene

A procedure is described for determining the mode and magnitude of gene-dosage compensation of transformed genes. It involves measurement of the ratio of the activity of a gene inserted at X-linked sites to the activity of the same gene inserted at autosomal sites. Applying the procedure to the Drosophila pseudoobscura Hsp82 gene inserted at ectopic sites in D. melanogaster and taking gene activity as proportional to the amount of transcript per gene copy, we conclude that (1) in both adults and larvae the gene is not compensated at autosomal sites or at a site in β-heterochromatin at the base of the X chromosome and is fully compensated at euchromatic X-chromosomal sites; (2) inappropriate normalization is responsible for a claim that the gene is compensated at autosomal sites; and (3) the observed compensation operates mainly or entirely by heightened activity of X-linked genes in males, rather than by reduced activity in females.

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Age-dependent ribosomal DNA variations and their effect on cellular function in mammalian cells

10.1101/2020.07.10.196840 ◽

2020 ◽

Author(s):

Eriko Watada ◽

Sihan Li ◽

Yutaro Hori ◽

Katsunori Fujiki ◽

Katsuhiko Shirahige ◽

...

Keyword(s):

Ribosomal Rna ◽

Copy Number ◽

Mammalian Cells ◽

Bone Marrow Cells ◽

Budding Yeast ◽

Rdna Repeat ◽

Age Dependent ◽

Old Mice ◽

Rdna Copy ◽

Rdna Copy Number

AbstractThe ribosomal RNA gene, which consists of tandem repetitive arrays (rDNA repeat), is one of the most unstable regions in the genome. The rDNA repeat in the budding yeast is known to become unstable as the cell ages. However, it is unclear how the rDNA repeat changes in ageing mammalian cells. Using quantitative analyses, we identified age-dependent alterations in rDNA copy number and levels of methylation in mice. The degree of methylation and copy number of rDNA from bone marrow cells of 2-year-old mice were increased by comparison to 4-week-old mice in two mouse strains, BALB/cA and C57BL/6. Moreover, the level of pre-rRNA transcripts was reduced in older BALB/cA mice. We also identified many sequence variations among the repeats with two mutations being unique to old mice. These sequences were conserved in budding yeast and equivalent mutations shortened the yeast chronological lifespan. Our findings suggest that rDNA is also fragile in mammalian cells and alterations within this region have a profound effect on cellular function.Author SummaryThe ribosomal RNA gene (rDNA) is one of the most unstable regions in the genome due to its tandem repetitive structure. rDNA copy number in the budding yeast increases and becomes unstable as the cell ages. It is speculated that the rDNA produces an “aging signal” inducing senescence and death. However, it is unclear how the rDNA repeat changes during the aging process in mammalian cells. In this study, we attempted to identify the age-dependent alteration of rDNA in mice. Using quantitative single cell analysis, we show that rDNA copy number increases in old mice bone marrow cells. By contrast, the level of ribosomal RNA production was reduced because of increased levels of DNA methylation that represses transcription. We also identified many sequence variations in the rDNA. Among them, three mutations were unique to old mice and two of them were found in the conserved region in budding yeast. We then established a yeast strain with the old mouse-specific mutations and found this shortened the lifespan of the cells. These findings suggest that rDNA is also fragile in mammalian cells and alteration to this region of the genome affects cellular senescence.

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