scholarly journals Complex Minisatellite Rearrangements Generated in the Total or Partial Absence of Rad27/hFEN1 Activity Occur in a Single Generation and Are Rad51 and Rad52 Dependent

2006 ◽  
Vol 26 (17) ◽  
pp. 6675-6689 ◽  
Author(s):  
Judith Lopes ◽  
Cyril Ribeyre ◽  
Alain Nicolas
Keyword(s):  
Pedigree Analysis ◽  
Model System ◽  
Yeast Cells ◽  
Single Generation ◽  
Daughter Cells ◽  
Double Deletion ◽  
Mother And Daughter ◽  
Complex Events ◽  
High Degree ◽  
Insight Into

ABSTRACT Genomes contain tandem repeat blocks that are at risk of expansion or contraction. The mechanisms of destabilization of the human minisatellite CEB1 (arrays of 36- to 43-bp repeats) were investigated in a previously developed model system, in which CEB1-0.6 (14 repeats) and CEB1-1.8 (42 repeats) alleles were inserted into the genome of Saccharomyces cerevisiae. As in human cells, CEB1 is stable in mitotically growing yeast cells but is frequently rearranged in the absence of the Rad27/hFEN1 protein involved in Okazaki fragments maturation. To gain insight into this mode of destabilization, the CEB1-1.8 and CEB1-0.6 human alleles and 47 rearrangements derived from a CEB1-1.8 progenitor in rad27Δ cells were sequenced. A high degree of polymorphism of CEB1 internal repeats was observed, attesting to a large variety of homology-driven rearrangements. Simple deletion, double deletion, and highly complex events were observed. Pedigree analysis showed that all rearrangements, even the most complex, occurred in a single generation and were inherited equally by mother and daughter cells. Finally, the rearrangement frequency was found to increase with array size, and partial complementation of the rad27Δ mutation by hFEN1 demonstrated that the production of novel CEB1 alleles is Rad52 and Rad51 dependent. Instability can be explained by an accumulation of unresolved flap structures during replication, leading to the formation of recombinogenic lesions and faulty repair, best understood by homology-dependent synthesis-strand displacement and annealing.

scholarly journals Hsp104 Overexpression Cures Saccharomyces cerevisiae [ PSI + ] by Causing Dissolution of the Prion Seeds

2014 ◽  
Vol 13 (5) ◽  
pp. 635-647 ◽  
Author(s):  
Yang-Nim Park ◽  
Xiaohong Zhao ◽  
Yang-In Yim ◽  
Horia Todor ◽  
Robyn Ellerbrock ◽  
...  
Keyword(s):  
Molecular Chaperone ◽  
Fluorescent Protein ◽  
Yeast Cells ◽  
Yeast Prion ◽  
Yeast Prions ◽  
Daughter Cells ◽  
Amyloid Aggregates ◽  
Mother And Daughter ◽  
Green Fluorescent ◽  
Kinetics Of

ABSTRACT The [ PSI + ] yeast prion is formed when Sup35 misfolds into amyloid aggregates. [ PSI + ], like other yeast prions, is dependent on the molecular chaperone Hsp104, which severs the prion seeds so that they pass on as the yeast cells divide. Surprisingly, however, overexpression of Hsp104 also cures [ PSI + ]. Several models have been proposed to explain this effect: inhibition of severing, asymmetric segregation of the seeds between mother and daughter cells, and dissolution of the prion seeds. First, we found that neither the kinetics of curing nor the heterogeneity in the distribution of the green fluorescent protein (GFP)-labeled Sup35 foci in partially cured yeast cells is compatible with Hsp104 overexpression curing [ PSI + ] by inhibiting severing. Second, we ruled out the asymmetric segregation model by showing that the extent of curing was essentially the same in mother and daughter cells and that the fluorescent foci did not distribute asymmetrically, but rather, there was marked loss of foci in both mother and daughter cells. These results suggest that Hsp104 overexpression cures [ PSI + ] by dissolution of the prion seeds in a two-step process. First, trimming of the prion seeds by Hsp104 reduces their size, and second, their amyloid core is eliminated, most likely by proteolysis.

Physiological Pharmacokinetic Models: Some Aspects of Theory, Practice and Potential

1988 ◽  
Vol 4 (2) ◽  
pp. 151-171 ◽  
Author(s):  
Richard W. D'souza ◽  
Harold Boxenbaum
Keyword(s):  
Circulatory System ◽  
Model System ◽  
Pharmacokinetic Models ◽  
Deep Insight ◽  
Pharmacokinetics And Pharmacodynamics ◽  
Pharmacodynamic Response ◽  
Physiological Models ◽  
And Behavior ◽  
High Degree ◽  
Insight Into

Models are intellectual constructs that pattern selected relationships among the elements of one system to correspond in some way to elements of a second system. In pharmacokinetics, physiological models provide a clearly articulated, rational, explanatory basis for the integration of empirical data; they do this by partitioning the biological system into relevant components (tissues, organs, etc.) and linking them together through the circulatory system. Unlike conventional mammillary compartment models, there is a clear correspondence between model system elements and physiological entities. By virtue of their high degree of physical and biochemical relevance, these models can help provide deep insight into structure, function and mechanism. Pharmacokinetic (and potentially pharmacodynamic) response-time relationships can thus be understood in terms of interconnections and behavior of constituent subsystems. At their worst, these models provide stale or infertile views of reality and thus frustrate and alienate us with the triviality of their insights. At their best, they allow us to understand the accumulation of thought in pharmacokinetics and pharmacodynamics, and help with the integration of data and improvement of experimental design.

scholarly journals Solid-phase inclusion as a mechanism for regulating unfolded proteins in the mitochondrial matrix

2020 ◽  
Vol 6 (32) ◽  
pp. eabc7288
Author(s):  
Linhao Ruan ◽  
Joshua T. McNamara ◽  
Xi Zhang ◽  
Alexander Chih-Chieh Chang ◽  
Jin Zhu ◽  
...  
Keyword(s):  
Solid Phase ◽  
Tricarboxylic Acid Cycle ◽  
Yeast Cells ◽  
Mitochondrial Proteins ◽  
Mitochondrial Matrix ◽  
Unfolded Proteins ◽  
Daughter Cells ◽  
Contact Sites ◽  
Age Dependent ◽  
Mother And Daughter

Proteostasis declines with age, characterized by the accumulation of unfolded or damaged proteins. Recent studies suggest that proteins constituting pathological inclusions in neurodegenerative diseases also enter and accumulate in mitochondria. How unfolded proteins are managed within mitochondria remains unclear. Here, we found that excessive unfolded proteins in the mitochondrial matrix of yeast cells are consolidated into solid-phase inclusions, which we term deposits of unfolded mitochondrial proteins (DUMP). Formation of DUMP occurs in mitochondria near endoplasmic reticulum–mitochondria contact sites and is regulated by mitochondrial proteins controlling the production of cytidine 5′-diphosphate–diacylglycerol. DUMP formation is age dependent but accelerated by exogenous unfolded proteins. Many enzymes of the tricarboxylic acid cycle were enriched in DUMP. During yeast cell division, DUMP formation is necessary for asymmetric inheritance of damaged mitochondrial proteins between mother and daughter cells. We provide evidence that DUMP-like structures may be induced by excessive unfolded proteins in human cells.

scholarly journals A novel factor essential for unconventional secretion of chitinase Cts1

2020 ◽  
Author(s):  
Michèle Reindl ◽  
Janpeter Stock ◽  
Kai P. Hussnaetter ◽  
Aycin Genc ◽  
Andreas Brachmann ◽  
...  
Keyword(s):  
Cell Division ◽  
Underlying Mechanism ◽  
Yeast Cells ◽  
Subcellular Targeting ◽  
Uv Mutagenesis ◽  
Daughter Cells ◽  
Secretion Pathway ◽  
Unconventional Secretion ◽  
Mother And Daughter ◽  
Two Hybrid

AbstractSubcellular targeting of proteins is essential to orchestrate cytokinesis in eukaryotic cells. During cell division of Ustilago maydis, for example, chitinases must be specifically targeted to the fragmentation zone at the site of cell division to degrade remnant chitin and thus separate mother and daughter cells. Chitinase Cts1 is exported to this location via an unconventional secretion pathway putatively operating in a lock-type manner. The underlying mechanism is largely unexplored. Here, we applied a forward genetic screen based on UV mutagenesis to identify components essential for Cts1 export. The screen revealed a novel factor termed Jps1 lacking known protein domains. Deletion of the corresponding gene confirmed its essential role for Cts1 secretion. Localization studies demonstrated that Jps1 colocalizes with Cts1 in the fragmentation zone of dividing yeast cells. While loss of Jps1 leads to exclusion of Cts1 from the fragmentation zone and strongly reduced unconventional secretion, deletion of the chitinase does not disturb Jps1 localization. Yeast-two hybrid experiments suggest that the two proteins interact. In essence, we identified a novel component of unconventional secretion that functions in the fragmentation zone to enable export of Cts1. We hypothesize that Jps1 acts as an anchoring factor, supporting the proposed novel lock-type mechanism of unconventional secretion.

Difference in generation times for mother and daughter cells in yeasts

1978 ◽  
Vol 24 (7) ◽  
pp. 827-833 ◽  
Author(s):  
T. W. Flegel
Keyword(s):  
Growth Characteristic ◽  
Time Lapse ◽  
Yeast Cells ◽  
Daughter Cells ◽  
Yeast Strains ◽  
Generation Times ◽  
Spot Check ◽  
Mother And Daughter ◽  
The Mean ◽  
Mother Cells

Fruitless attempts to synchronize haploid yeast cells from the Phragmobasidiomycete Sirobasidium magnum led to the discovery that the mother and daughter cells (MDC) had greatly different generation times. Time-lapse photographic sequences of budding showed that the mean generation time for daughter cells was more than three times greater than that for mother cells. This growth characteristic could be determined by a spot check of the microcolony pattern on agar. Using such a check, yeast strains of Rhodotorula (Rhodosporidium) and Cryptococcus that were tested demonstrated relative MDC equivalence while those of Sporobolomyces, Bullera, and Tremella showed MDC non-equivalence in varying degrees.

scholarly journals Age-induced P-bodies become detrimental and shorten the lifespan of yeast

2021 ◽  
Author(s):  
Joonhyuk Choi ◽  
Shuhao Wang ◽  
Yang Li ◽  
Nan Hao ◽  
Brian M Zid
Keyword(s):  
Irreversible Process ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Model System ◽  
Yeast Cells ◽  
P Bodies ◽  
Daughter Cells ◽  
Progressive Loss ◽  
External Stresses ◽  
Mother Cells

Aging is an irreversible process characterized by a progressive loss of homeostasis in cells, which often manifests as protein aggregates. Recently, it has been speculated that aggregates of RNA-binding proteins (RBPs) may go through pathological transitions during aging and drive the progression of age-associated neurodegenerative diseases. Using Saccharomyces cerevisiae as a model system of aging, we find that P-bodies - an RBP granule that is formed and can be beneficial for cell growth during stress conditions - naturally form during aging without any external stresses and an increase in P-body intensity is negatively correlated with the future lifespan of yeast cells. When mother cells transfer age-induced P-bodies to daughter cells, the mother cells extend lifespan, while the daughter cells grow poorly, suggesting that these age-induced P-bodies may be directly pathological. Furthermore, we find that suppressing acidification of the cytosol during aging slows down the increase in the intensity of P-body foci and extends lifespan. Our data suggest that acidification of the cytosol may facilitate the pathological transition of RBP granules during aging.

scholarly journals Rate volatility and asymmetric segregation diversify mutation burden in mutator cells

2020 ◽  
Author(s):  
I.T. Dowsett ◽  
J. Sneeden ◽  
B.J. Olson ◽  
J. McKay-Fleisch ◽  
E. McAuley ◽  
...  
Keyword(s):  
Mismatch Repair ◽  
Dna Polymerase ◽  
Count Data ◽  
Yeast Cells ◽  
Cancer Evolution ◽  
Mendelian Segregation ◽  
Replication Errors ◽  
Daughter Cells ◽  
Mutation Burden ◽  
Mother And Daughter

Mutations that compromise mismatch repair (MMR) or DNA polymerase exonuclease domains produce mutator phenotypes capable of fueling cancer evolution. Tandem defects in these pathways dramatically increase mutation rate. Here, we model how mutator phenotypes expand genetic heterogeneity in budding yeast cells using a single-cell resolution approach that tallies all replication errors arising from individual divisions. The distribution of count data from cells lacking MMR and polymerase proofreading was broader than expected for a single rate, consistent with volatility of the mutator phenotype. The number of mismatches that segregated to the mother and daughter cells after the initial round of replication co-varied, suggesting that mutagenesis in each division is governed by a different underlying rate. The distribution of “fixed” mutation counts that cells inherit is further broadened by an unequal sharing of mutations due to semiconservative replication and Mendelian segregation. Modeling suggests that this asymmetric segregation may diversify mutation burden in mutator-driven tumors.

scholarly journals Peroxisome reintroduction in Hansenula polymorpha requires Pex25 and Rho1

2011 ◽  
Vol 193 (5) ◽  
pp. 885-900 ◽  
Author(s):  
Ruchi Saraya ◽  
Arjen M. Krikken ◽  
Marten Veenhuis ◽  
Ida J. van der Klei
Keyword(s):  
Mutant Strain ◽  
De Novo ◽  
Hansenula Polymorpha ◽  
Protein Family ◽  
Yeast Cells ◽  
Deficient Mutant ◽  
Wild Type ◽  
Double Deletion ◽  
Insight Into

We identified two proteins, Pex25 and Rho1, which are involved in reintroduction of peroxisomes in peroxisome-deficient yeast cells. These are, together with Pex3, the first proteins identified as essential for this process. Of the three members of the Hansenula polymorpha Pex11 protein family—Pex11, Pex25, and Pex11C—only Pex25 was required for reintroduction of peroxisomes into a peroxisome-deficient mutant strain. In peroxisome-deficient pex3 cells, Pex25 localized to structures adjacent to the ER, whereas in wild-type cells it localized to peroxisomes. Pex25 cells were not themselves peroxisome deficient but instead contained a slightly increased number of peroxisomes. Interestingly, pex11 pex25 double deletion cells, in which both peroxisome fission (due to the deletion of PEX11) and reintroduction (due to deletion of PEX25) was blocked, did display a peroxisome-deficient phenotype. Peroxisomes reappeared in pex11 pex25 cells upon synthesis of Pex25, but not of Pex11. Reintroduction in the presence of Pex25 required the function of the GTPase Rho1. These data therefore provide new and detailed insight into factors important for de novo peroxisome formation in yeast.

scholarly journals A Mutation in the ATP2 Gene Abrogates the Age Asymmetry Between Mother and Daughter Cells of the Yeast Saccharomyces cerevisiae

Genetics ◽  
2002 ◽  
Vol 162 (1) ◽  
pp. 73-87 ◽  
Author(s):  
Chi-Yung Lai ◽  
Ewa Jaruga ◽  
Corina Borghouts ◽  
S Michal Jazwinski
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Division ◽  
Life Span ◽  
Mother Cell ◽  
Yeast Cells ◽  
Mitochondrial Morphology ◽  
Temperature Sensitive ◽  
Yeast Saccharomyces Cerevisiae ◽  
Daughter Cells ◽  
Mother And Daughter

Abstract The yeast Saccharomyces cerevisiae reproduces by asymmetric cell division, or budding. In each cell division, the daughter cell is usually smaller and younger than the mother cell, as defined by the number of divisions it can potentially complete before it dies. Although individual yeast cells have a limited life span, this age asymmetry between mother and daughter ensures that the yeast strain remains immortal. To understand the mechanisms underlying age asymmetry, we have isolated temperature-sensitive mutants that have limited growth capacity. One of these clonal-senescence mutants was in ATP2, the gene encoding the β-subunit of mitochondrial F1, F0-ATPase. A point mutation in this gene caused a valine-to-isoleucine substitution at the ninetieth amino acid of the mature polypeptide. This mutation did not affect the growth rate on a nonfermentable carbon source. Life-span determinations following temperature shift-down showed that the clonal-senescence phenotype results from a loss of age asymmetry at 36°, such that daughters are born old. It was characterized by a loss of mitochondrial membrane potential followed by the lack of proper segregation of active mitochondria to daughter cells. This was associated with a change in mitochondrial morphology and distribution in the mother cell and ultimately resulted in the generation of cells totally lacking mitochondria. The results indicate that segregation of active mitochondria to daughter cells is important for maintenance of age asymmetry and raise the possibility that mitochondrial dysfunction may be a normal cause of aging. The finding that dysfunctional mitochondria accumulated in yeasts as they aged and the propensity for old mother cells to produce daughters depleted of active mitochondria lend support to this notion. We propose, more generally, that age asymmetry depends on partition of active and undamaged cellular components to the progeny and that this “filter” breaks down with age.

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