Robertsonian chromosomes form by fusion of two chromosomes that have centromeres located near their ends, known as acrocentric or telocentric chromosomes. This fusion creates a new metacentric ...chromosome and is a major mechanism of karyotype evolution and speciation. Robertsonian chromosomes are common in nature and were first described in grasshoppers by the zoologist W. R. B. Robertson more than 100 years ago. They have since been observed in many species, including catfish, sheep, butterflies, bats, bovids, rodents and humans, and are the most common chromosomal change in mammals. Robertsonian translocations are particularly rampant in the house mouse, Mus musculus domesticus, where they exhibit meiotic drive and create reproductive isolation. Recent progress has been made in understanding how Robertsonian chromosomes form in the human genome, highlighting some of the fundamental principles of how and why these types of fusion events occur so frequently. Consequences of these fusions include infertility and Down's syndrome. In this Hypothesis, I postulate that the conditions that allow these fusions to form are threefold: (1) sequence homology on non-homologous chromosomes, often in the form of repetitive DNA; (2) recombination initiation during meiosis; and (3) physical proximity of the homologous sequences in three-dimensional space. This Hypothesis highlights the latest progress in understanding human Robertsonian translocations within the context of the broader literature on Robertsonian chromosomes.
Ribosomal RNAs (rRNAs) in budding yeast are encoded by ~100-200 repeats of a 9.1kb sequence arranged in tandem on chromosome XII, the ribosomal DNA (rDNA) locus. Copy number of rDNA repeat units in ...eukaryotic cells is maintained far in excess of the requirement for ribosome biogenesis. Despite the importance of the repeats for both ribosomal and non-ribosomal functions, it is currently not known how "normal" copy number is determined or maintained. To identify essential genes involved in the maintenance of rDNA copy number, we developed a droplet digital PCR based assay to measure rDNA copy number in yeast and used it to screen a yeast conditional temperature-sensitive mutant collection of essential genes. Our screen revealed that low rDNA copy number is associated with compromised DNA replication. Further, subculturing yeast under two separate conditions of DNA replication stress selected for a contraction of the rDNA array independent of the replication fork blocking protein, Fob1. Interestingly, cells with a contracted array grew better than their counterparts with normal copy number under conditions of DNA replication stress. Our data indicate that DNA replication stresses select for a smaller rDNA array. We speculate that this liberates scarce replication factors for use by the rest of the genome, which in turn helps cells complete DNA replication and continue to propagate. Interestingly, tumors from mini chromosome maintenance 2 (MCM2)-deficient mice also show a loss of rDNA repeats. Our data suggest that a reduction in rDNA copy number may indicate a history of DNA replication stress, and that rDNA array size could serve as a diagnostic marker for replication stress. Taken together, these data begin to suggest the selective pressures that combine to yield a "normal" rDNA copy number.
Ribosomal DNA is one of the most variable regions in the human genome with respect to copy number. Despite the importance of rDNA for cellular function, we know virtually nothing about what governs ...its copy number, stability, and sequence in the mammalian genome due to challenges associated with mapping and analysis. We applied computational and droplet digital PCR approaches to measure rDNA copy number in normal and cancer states in human and mouse genomes. We find that copy number and sequence can change in cancer genomes. Counterintuitively, human cancer genomes show a loss of copies, accompanied by global copy number co-variation. The sequence can also be more variable in the cancer genome. Cancer genomes with lower copies have mutational evidence of mTOR hyperactivity. The PTEN phosphatase is a tumor suppressor that is critical for genome stability and a negative regulator of the mTOR kinase pathway. Surprisingly, but consistent with the human cancer genomes, hematopoietic cancer stem cells from a Pten-/- mouse model for leukemia have lower rDNA copy number than normal tissue, despite increased proliferation, rRNA production, and protein synthesis. Loss of copies occurs early and is associated with hypersensitivity to DNA damage. Therefore, copy loss is a recurrent feature in cancers associated with mTOR activation. Ribosomal DNA copy number may be a simple and useful indicator of whether a cancer will be sensitive to DNA damaging treatments.
The nucleolus constitutes a prominent nuclear compartment, a membraneless organelle that was first documented in the 1830s. The fact that specific chromosomal regions were present in the nucleolus ...was recognized by Barbara McClintock in the 1930s, and these regions were termed nucleolar organizing regions, or NORs. The primary function of ribosomal DNA (rDNA) is to produce RNA components of ribosomes. Yet, ribosomal DNA also plays a pivotal role in nuclear organization by assembling the nucleolus. This review is focused on the rDNA and associated proteins in the context of genome organization. Recent advances in understanding chromatin organization suggest that chromosomes are organized into topological domains by a DNA loop extrusion process. We discuss the perspective that rDNA may also be organized in topological domains constrained by structural maintenance of chromosome protein complexes such as cohesin and condensin. Moreover, biophysical studies indicate that the nucleolar compartment may be formed by active processes as well as phase separation, a perspective that lends further insight into nucleolar organization. The application of the latest perspectives and technologies to this organelle help further elucidate its role in nuclear structure and function.
Structural maintenance of chromosome (SMC) protein complexes, including cohesin and condensin, are increasingly being recognized for their important role in cancer and development, making it critical ...that we understand how these evolutionarily conserved multi-subunit protein complexes associate with and organize the genome. We review adaptor proteins for SMC complexes and how these adaptors may capture SMC complexes following loop extrusion to provide a framework for chromosome organization.
Ribosomes are large, multi-subunit ribonucleoprotein complexes, essential for protein synthesis. To meet the high cellular demand for ribosomes, all eukaryotes have numerous copies of ribosomal DNA ...(rDNA) genes that encode ribosomal RNA (rRNA), usually far in excess of the requirement for ribosome biogenesis. In all eukaryotes studied, rDNA genes are arranged in one or more clusters of tandem repeats localized to nucleoli. The tandem arrangement of repeats, combined with the high rates of transcription at the rDNA loci, and the difficulty of replicating repetitive sequences make the rDNA inherently unstable and particularly susceptible to large variations in repeat copy number. Despite mounting evidence suggesting extra-ribosomal functions of the rDNA, its repetitive nature has excluded it from traditional sequencing-based studies. However, more recently, several studies have revealed the unique potential of the rDNA to act as a “canary in the coalmine,” being particularly sensitive to genomic stresses and acting as a source of adaptive response. Here, we review evidence uncovering mechanisms of regulation of instability and copy number variation at the rDNA and their role in adaptation to the environment, which could serve to understand the basic principles governing the behavior of other tandem repeats and their role in shaping the genome.
Summary
Reproductive aging is characterized by a marked decline in oocyte quality that contributes to infertility, miscarriages, and birth defects. This decline is multifactorial, and the underlying ...mechanisms are under active investigation. Here, we performed RNA‐Seq on individual growing follicles from reproductively young and old mice to identify age‐dependent functions in oocytes. This unbiased approach revealed genes involved in cellular processes known to change with age, including mitochondrial function and meiotic chromosome segregation, but also uncovered previously unappreciated categories of genes related to proteostasis and organelles required for protein metabolism. We further validated our RNA‐Seq data by comparing nucleolar structure and function in oocytes from reproductively young and old mice, as this organelle is central for protein production. We examined key nucleolar markers, including upstream binding transcription factor (UBTF), an RNA polymerase I cofactor, and fibrillarin, an rRNA methyltransferase. In oocytes from mice of advanced reproductive age, UBTF was primarily expressed in giant fibrillar centers (GFCs), structures associated with high levels of rDNA transcription, and fibrillarin expression was increased ~2‐fold. At the ultrastructural level, oocyte nucleoli from reproductively old mice had correspondingly more prominent fibrillar centers and dense fibrillar centers relative to young controls and more ribosomes were found in the cytoplasm. Taken together, our findings are significant because the growing oocyte is one of the most translationally active cells in the body and must accumulate high‐quality maternally derived proteins to support subsequent embryo development. Thus, perturbations in protein metabolism are likely to have a profound impact on gamete health.
Nuclear pore complexes play central roles as gatekeepers of RNA and protein transport between the cytoplasm and nucleoplasm. However, their large size and dynamic nature have impeded a full ...structural and functional elucidation. Here we determined the structure of the entire 552-protein nuclear pore complex of the yeast Saccharomyces cerevisiae at sub-nanometre precision by satisfying a wide range of data relating to the molecular arrangement of its constituents. The nuclear pore complex incorporates sturdy diagonal columns and connector cables attached to these columns, imbuing the structure with strength and flexibility. These cables also tie together all other elements of the nuclear pore complex, including membrane-interacting regions, outer rings and RNA-processing platforms. Inwardly directed anchors create a high density of transport factor-docking Phe-Gly repeats in the central channel, organized into distinct functional units. This integrative structure enables us to rationalize the architecture, transport mechanism and evolutionary origins of the nuclear pore complex.
Cohesin is an evolutionarily conserved chromosome-associated protein complex essential for chromosome segregation, gene expression, and repair of DNA damage. Mutations that affect this complex cause ...the human developmental disorder Cornelia de Lange syndrome (CdLS), thought to arise from defective embryonic transcription. We establish a significant role for placental defects in the development of CdLS mouse embryos (Nipbl and Hdac8). Placenta is a naturally senescent tissue; we demonstrate that persistent DNA damage potentiates senescence and activates cytokine signaling. Mutant embryo developmental outcomes are significantly improved in the context of a wild-type placenta or by genetically restricting cytokine signaling. Our study highlights that cohesin is required for maintaining ploidy and the repair of spontaneous DNA damage in placental cells, suggesting that genotoxic stress and ensuing placental senescence and cytokine production could represent a broad theme in embryo health and viability.
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•CdLS mouse models show placental defects•Impaired cohesin function increases ploidy, DNA damage, and senescence in the placenta•Blocking cytokine signaling in the placenta improves CdLS embryo outcomes•Genotoxic stress in the placenta negatively impacts embryo outcomes
Using mouse models for Cornelia de Lange syndrome with impaired cohesin function, Singh et al. demonstrate that persistent DNA damage in placental cells is associated with placental senescence and cytokine secretion with negative impacts on embryo outcomes. Genotoxic stress and senescence in placenta may generally perturb embryo development and viability.
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