scholarly journals The silencing factor Sir3 is a molecular bridge that sticks together distant loci

2020 ◽  
Author(s):  
Myriam Ruault ◽  
Vittore F. Scolari ◽  
Luciana Lazar-Stefanita ◽  
Antoine Hocher ◽  
Isabelle Loïodice ◽  
...  
Keyword(s):  
Long Range ◽  
Molecular Mechanisms ◽  
Budding Yeast ◽  
Synthetic Approach ◽  
Molecular Bridge ◽  
Subtelomeric Regions

ABSTRACTPhysical contacts between distant loci contribute to regulate genome function. However, the molecular mechanisms responsible for settling and maintaining such interactions remain poorly understood. Here we investigate the well conserved interactions between heterochromatin loci. In budding yeast, the 32 telomeres cluster in 3-5 foci in exponentially growing cells. This clustering is functionally linked to the formation of heterochromatin in subtelomeric regions through the recruitment of the silencing complex SIR composed of Sir2/3/4. Combining microscopy and Hi-C on strains expressing different alleles of SIR3, we show that the binding of Sir3 directly promotes long range contacts between distant regions, including the rDNA, telomeres, and internal Sir3 bound sites. Furthermore, we unveil a new property of Sir3 in promoting rDNA compaction. Finally, using a synthetic approach we demonstrate that Sir3 can bond loci belonging to different chromosomes together, when targeted to these loci, independently of its interaction with its known partners (Rap1, and Sir4), Sir2 activity or chromosome context. Altogether these data suggest that Sir3 represents an uncommon example of protein able to bridge directly distant loci.

scholarly journals Clustering heterochromatin: Sir3 promotes telomere clustering independently of silencing in yeast

2011 ◽  
Vol 192 (3) ◽  
pp. 417-431 ◽  
Author(s):  
Myriam Ruault ◽  
Arnaud De Meyer ◽  
Isabelle Loïodice ◽  
Angela Taddei
Keyword(s):  
Long Range ◽  
Binding Sites ◽  
General Feature ◽  
Deoxyribonucleic Acid ◽  
Budding Yeast ◽  
Nuclear Interior ◽  
Long Range Interactions ◽  
Subtelomeric Regions

A general feature of the nucleus is the organization of repetitive deoxyribonucleic acid sequences in clusters concentrating silencing factors. In budding yeast, we investigated how telomeres cluster in perinuclear foci associated with the silencing complex Sir2–Sir3–Sir4 and found that Sir3 is limiting for telomere clustering. Sir3 overexpression triggers the grouping of telomeric foci into larger foci that relocalize to the nuclear interior and correlate with more stable silencing in subtelomeric regions. Furthermore, we show that Sir3′s ability to mediate telomere clustering can be separated from its role in silencing. Indeed, nonacetylable Sir3, which is unable to spread into subtelomeric regions, can mediate telomere clustering independently of Sir2–Sir4 as long as it is targeted to telomeres by the Rap1 protein. Thus, arrays of Sir3 binding sites at telomeres appeared as the sole requirement to promote trans-interactions between telomeres. We propose that similar mechanisms involving proteins able to oligomerize account for long-range interactions that impact genomic functions in many organisms.

scholarly journals CpG island mapping of a mouse double-minute chromosome.

1993 ◽  
Vol 13 (8) ◽  
pp. 4459-4464 ◽  
Author(s):  
J L Beland ◽  
J A Longo ◽  
P J Hahn
Keyword(s):  
Cell Line ◽  
Long Range ◽  
Molecular Mechanisms ◽  
Cpg Island ◽  
Single Copy ◽  
Restriction Map ◽  
Chromosomal Dna ◽  
Dhfr Gene ◽  
Range Restriction ◽  
Double Minute

The development of double-minute chromosomes (DMs) and subsequent gene amplification are important genomic alterations resulting in increased oncogene expression in a variety of tumors. The molecular mechanisms mediating the development of these acentric extrachromosomal elements have not been completely defined. To elucidate the mechanisms involved in DM formation, we have developed strategies to map amplified circular DM DNA. In this study, we present a long-range restriction map of a 980-kb DM. A cell line cloned from mouse EMT-6 cells was developed by stepwise selection for resistance to methotrexate. This cloned cell line contains multiple copies of the 980-kb DM carrying the dihydrofolate reductase (DHFR) gene. A long-range restriction map was developed in which a hypomethylated CpG-rich region near the DHFR gene served as a landmark. This strategy was combined with plasmid-like analysis of ethidium bromide-stained pulsed-field gels and indicated that a single copy of the DHFR gene was located near a hypomethylated region containing SsII and NotI sites. At least 490 kb of this DM appears to be composed of unrearranged chromosomal DNA.

CpG island mapping of a mouse double-minute chromosome

1993 ◽  
Vol 13 (8) ◽  
pp. 4459-4464
Author(s):  
J L Beland ◽  
J A Longo ◽  
P J Hahn
Keyword(s):  
Cell Line ◽  
Long Range ◽  
Molecular Mechanisms ◽  
Cpg Island ◽  
Single Copy ◽  
Restriction Map ◽  
Chromosomal Dna ◽  
Dhfr Gene ◽  
Range Restriction ◽  
Double Minute

The development of double-minute chromosomes (DMs) and subsequent gene amplification are important genomic alterations resulting in increased oncogene expression in a variety of tumors. The molecular mechanisms mediating the development of these acentric extrachromosomal elements have not been completely defined. To elucidate the mechanisms involved in DM formation, we have developed strategies to map amplified circular DM DNA. In this study, we present a long-range restriction map of a 980-kb DM. A cell line cloned from mouse EMT-6 cells was developed by stepwise selection for resistance to methotrexate. This cloned cell line contains multiple copies of the 980-kb DM carrying the dihydrofolate reductase (DHFR) gene. A long-range restriction map was developed in which a hypomethylated CpG-rich region near the DHFR gene served as a landmark. This strategy was combined with plasmid-like analysis of ethidium bromide-stained pulsed-field gels and indicated that a single copy of the DHFR gene was located near a hypomethylated region containing SsII and NotI sites. At least 490 kb of this DM appears to be composed of unrearranged chromosomal DNA.

scholarly journals Molecular mechanisms facilitating the initial kinetochore encounter with spindle microtubules

2017 ◽  
Vol 216 (6) ◽  
pp. 1609-1622 ◽  
Author(s):  
Vanya Vasileva ◽  
Marek Gierlinski ◽  
Zuojun Yue ◽  
Nicola O’Reilly ◽  
Etsushi Kitamura ◽  
...  
Keyword(s):  
Half Life ◽  
Molecular Mechanisms ◽  
Significant Contribution ◽  
Budding Yeast ◽  
Computational Simulation ◽  
Spindle Microtubule ◽  
Multiple Factors ◽  
Spindle Microtubules ◽  
Rate Limiting

The initial kinetochore (KT) encounter with a spindle microtubule (MT; KT capture) is one of the rate-limiting steps in establishing proper KT–MT interaction during mitosis. KT capture is facilitated by multiple factors, such as MT extension in various directions, KT diffusion, and MT pivoting. In addition, KTs generate short MTs, which subsequently interact with a spindle MT. KT-derived MTs may facilitate KT capture, but their contribution is elusive. In this study, we find that Stu1 recruits Stu2 to budding yeast KTs, which promotes MT generation there. By removing Stu2 specifically from KTs, we show that KT-derived MTs shorten the half-life of noncaptured KTs from 48–49 s to 28–34 s. Using computational simulation, we found that multiple factors facilitate KT capture redundantly or synergistically. In particular, KT-derived MTs play important roles both by making a significant contribution on their own and by synergistically enhancing the effects of KT diffusion and MT pivoting. Our study reveals fundamental mechanisms facilitating the initial KT encounter with spindle MTs.

scholarly journals Yeast as a Model to Understand Actin-Mediated Cellular Functions in Mammals—Illustrated with Four Actin Cytoskeleton Proteins

Cells ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 672 ◽  
Author(s):  
Zain Akram ◽  
Ishtiaq Ahmed ◽  
Heike Mack ◽  
Ramandeep Kaur ◽  
Richard C. Silva ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Molecular Mechanisms ◽  
Budding Yeast ◽  
Animal Cell ◽  
Molecular Genetic ◽  
In Vivo Studies ◽  
Yeast Saccharomyces Cerevisiae ◽  
Higher Eukaryotes

The budding yeast Saccharomyces cerevisiae has an actin cytoskeleton that comprises a set of protein components analogous to those found in the actin cytoskeletons of higher eukaryotes. Furthermore, the actin cytoskeletons of S. cerevisiae and of higher eukaryotes have some similar physiological roles. The genetic tractability of budding yeast and the availability of a stable haploid cell type facilitates the application of molecular genetic approaches to assign functions to the various actin cytoskeleton components. This has provided information that is in general complementary to that provided by studies of the equivalent proteins of higher eukaryotes and hence has enabled a more complete view of the role of these proteins. Several human functional homologues of yeast actin effectors are implicated in diseases. A better understanding of the molecular mechanisms underpinning the functions of these proteins is critical to develop improved therapeutic strategies. In this article we chose as examples four evolutionarily conserved proteins that associate with the actin cytoskeleton: (1) yeast Hof1p/mammalian PSTPIP1, (2) yeast Rvs167p/mammalian BIN1, (3) yeast eEF1A/eEF1A1 and eEF1A2 and (4) yeast Yih1p/mammalian IMPACT. We compare the knowledge on the functions of these actin cytoskeleton-associated proteins that has arisen from studies of their homologues in yeast with information that has been obtained from in vivo studies using live animals or in vitro studies using cultured animal cell lines.

scholarly journals A long-range chromatin interaction regulates SATB homeobox 1 gene expression in trophoblast stem cells

2020 ◽  
Author(s):  
Wei Yu ◽  
V. Praveen Chakravarthi ◽  
Shaon Borosha ◽  
Anamika Ratri ◽  
Khyati Dalal ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Polymerase Ii ◽  
Long Range ◽  
Molecular Mechanisms ◽  
Regulatory Region ◽  
Chromatin Interaction ◽  
Enhancer Activity ◽  
Histone Marks ◽  
Developmental Gene Expression ◽  
Trophoblast Stem

ABSTRACTSATB homeobox proteins are important regulators of developmental gene expression. Among the stem cell lineages determined during early embryonic development, trophoblast stem (TS) cells exhibit robust SATB expression. Both SATB1 and SATB2 act to maintain trophoblast stem-state. However, the molecular mechanisms that regulate TS-specific Satb expression are not yet known. We identified Satb1 variant 2 as the predominant transcript in trophoblasts. Histone marks, and RNA polymerase II occupancy in TS cells indicated active state of the promoter. A novel cis-regulatory region with active histone marks was identified ∼21kbp upstream of variant 2 promoter. CRISPR/Cas9 mediated disruption of this sequence decreased Satb1 expression in TS cells and chromatin conformation capture confirmed looping of this regulatory region into the promoter. Scanning position weight matrices across the enhancer predicted two ELF5 binding sites in close vicinity of SATB1 sites, which were confirmed by chromatin immunoprecipitation. Knockdown of ELF5 downregulated Satb1 expression in TS cells and overexpression of ELF5 increased the enhancer-reporter activity. Interestingly, ELF5 interacts with SATB1 in TS cells, and the enhancer activity was upregulated following SATB overexpression. Our findings indicate that trophoblast-specific Satb1 expression is regulated by long-range chromatin looping of an enhancer that interacts with ELF5 and SATB proteins.

scholarly journals Sir3 mediates long-range chromosome interactions in budding yeast

2021 ◽  
Vol 31 (3) ◽  
pp. 411-425 ◽  
Author(s):  
Myriam Ruault ◽  
Vittore F. Scolari ◽  
Luciana Lazar-Stefanita ◽  
Antoine Hocher ◽  
Isabelle Loïodice ◽  
...  
Keyword(s):  
Long Range ◽  
Budding Yeast

scholarly journals Genomic Instability and Cellular Senescence: Lessons From the Budding Yeast

2021 ◽  
Vol 8 ◽  
Author(s):  
Jee Whu Lee ◽  
Eugene Boon Beng Ong
Keyword(s):  
Ribosomal Dna ◽  
Cellular Senescence ◽  
Molecular Mechanisms ◽  
Budding Yeast ◽  
Genetic Alterations ◽  
Lessons Learned ◽  
Complex Biological Process ◽  
Dna Instability ◽  
Permanent Cell ◽  
Living Organisms

Aging is a complex biological process that occurs in all living organisms. Aging is initiated by the gradual accumulation of biomolecular damage in cells leading to the loss of cellular function and ultimately death. Cellular senescence is one such pathway that leads to aging. The accumulation of nucleic acid damage and genetic alterations that activate permanent cell-cycle arrest triggers the process of senescence. Cellular senescence can result from telomere erosion and ribosomal DNA instability. In this review, we summarize the molecular mechanisms of telomere length homeostasis and ribosomal DNA stability, and describe how these mechanisms are linked to cellular senescence and longevity through lessons learned from budding yeast.

scholarly journals The class V myosin motor protein, Myo2, plays a major role in mitochondrial motility in Saccharomyces cerevisiae

2008 ◽  
Vol 181 (1) ◽  
pp. 119-130 ◽  
Author(s):  
Katrin Altmann ◽  
Martina Frank ◽  
Daniel Neumann ◽  
Stefan Jakobs ◽  
Benedikt Westermann
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Actin Cytoskeleton ◽  
Molecular Mechanisms ◽  
Budding Yeast ◽  
Motor Protein ◽  
Mitochondrial Morphology ◽  
Direct Role ◽  
Class V ◽  
Myosin Motor ◽  
Mitochondrial Motility

The actin cytoskeleton is essential for polarized, bud-directed movement of cellular membranes in Saccharomyces cerevisiae and thus ensures accurate inheritance of organelles during cell division. Also, mitochondrial distribution and inheritance depend on the actin cytoskeleton, though the precise molecular mechanisms are unknown. Here, we establish the class V myosin motor protein, Myo2, as an important mediator of mitochondrial motility in budding yeast. We found that mutants with abnormal expression levels of Myo2 or its associated light chain, Mlc1, exhibit aberrant mitochondrial morphology and loss of mitochondrial DNA. Specific mutations in the globular tail of Myo2 lead to aggregation of mitochondria in the mother cell. Isolated mitochondria lacking functional Myo2 are severely impaired in their capacity to bind to actin filaments in vitro. Time-resolved fluorescence microscopy revealed a block of bud-directed anterograde mitochondrial movement in cargo binding–defective myo2 mutant cells. We conclude that Myo2 plays an important and direct role for mitochondrial motility and inheritance in budding yeast.

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