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 Peroxisome Maintenance Depends on De Novo Peroxisome Formation in Yeast Mutants Defective in Peroxisome Fission and Inheritance

2019 ◽  
Vol 20 (16) ◽  
pp. 4023 ◽  
Author(s):  
Justyna P. Wróblewska ◽  
Ida J. van der Klei
Keyword(s):  
De Novo ◽  
Hansenula Polymorpha ◽  
Yeast Cells ◽  
Wild Type ◽  
Double Deletion ◽  
Mother Cells ◽  
Yeast Mutants ◽  
Rescue Mechanism ◽  
Wild Type Yeast ◽  
In Cells

There is an ongoing debate on how peroxisomes form: by growth and fission of pre-existing peroxisomes or de novo from another membrane. It has been proposed that, in wild type yeast cells, peroxisome fission and careful segregation of the organelles over mother cells and buds is essential for organelle maintenance. Using live cell imaging we observed that cells of the yeast Hansenula polymorpha, lacking the peroxisome fission protein Pex11, still show peroxisome fission and inheritance. Also, in cells of mutants without the peroxisome inheritance protein Inp2 peroxisome segregation can still occur. In contrast, peroxisome fission and inheritance were not observed in cells of a pex11 inp2 double deletion strain. In buds of cells of this double mutant, new organelles likely appear de novo. Growth of pex11 inp2 cells on methanol, a growth substrate that requires functional peroxisomes, is retarded relative to the wild type control. Based on these observations we conclude that in H. polymorpha de novo peroxisome formation is a rescue mechanism, which is less efficient than organelle fission and inheritance to maintain functional peroxisomes.

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 Inhibition of Fungal Colonization by Pseudoalteromonas tunicata Provides a Competitive Advantage during Surface Colonization

2006 ◽  
Vol 72 (9) ◽  
pp. 6079-6087 ◽  
Author(s):  
A. Franks ◽  
S. Egan ◽  
C. Holmstr�m ◽  
S. James ◽  
H. Lappin-Scott ◽  
...  
Keyword(s):  
Competitive Advantage ◽  
Fungal Community ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Yeast Cells ◽  
Fungal Colonization ◽  
Antifungal Compound ◽  
Deficient Mutant ◽  
Wild Type ◽  
Surface Colonization ◽  
Pseudoalteromonas Tunicata

ABSTRACT The marine epiphytic bacterium Pseudoalteromonas tunicata produces a range of extracellular secondary metabolites that inhibit an array of common fouling organisms, including fungi. In this study, we test the hypothesis that the ability to inhibit fungi provides P. tunicata with an advantage during colonization of a surface. Studies on a transposon-generated antifungal-deficient mutant of P. tunicata, FM3, indicated that a long-chain fatty acid-coenzyme A ligase is involved in the production of a broad-range antifungal compound by P. tunicata. Flow cell experiments demonstrated that production of an antifungal compound provided P. tunicata with a competitive advantage against a marine yeast isolate during surface colonization. This compound enabled P. tunicata to disrupt an already established fungal biofilm by decreasing the number of yeast cells attached to the surface by 66% � 9%. For in vivo experiments, the wild-type and FM3 strains of P. tunicata were used to inoculate the surface of the green alga Ulva australis. Double-gradient denaturing gradient gel electrophoresis analysis revealed that after 48 h, the wild-type P. tunicata had outcompeted the surface-associated fungal community, whereas the antifungal-deficient mutant had no effect on the fungal community. Our data suggest that P. tunicata is an effective competitor against fungal surface communities in the marine environment.

scholarly journals A New Transformation-Deficient Mutant ofHaemophilus influenzae Rd with Normal DNA Uptake

1998 ◽  
Vol 180 (3) ◽  
pp. 746-748 ◽  
Author(s):  
M. L. Gwinn ◽  
R. Ramanathan ◽  
H. O. Smith ◽  
J.-F. Tomb
Keyword(s):  
Haemophilus Influenzae ◽  
Mutant Strain ◽  
Competence Development ◽  
Transformation Frequency ◽  
Dna Uptake ◽  
Deficient Mutant ◽  
Wild Type ◽  
Gram Negative ◽  
Ofhaemophilus Influenzae

ABSTRACT Haemophilus influenzae Rd is a gram-negative natural transformer. A mutant strain, RJ248, that has normal DNA uptake and translocation but whose transformation frequency is 300 times lower than that of wild-type H. influenzae and whose phage recombination is 8 times lower was isolated. The affected gene,comM, is induced during competence development in wild-typeH. influenzae but not in RJ248.

scholarly journals The Impact of Broadly Relevant Novel Discoveries on Student Project Ownership in a Traditional Lab Course Turned CURE

2019 ◽  
Vol 18 (4) ◽  
pp. ar57 ◽  
Author(s):  
Katelyn M. Cooper ◽  
Joseph N. Blattman ◽  
Taija Hendrix ◽  
Sara E. Brownell
Keyword(s):  
Immune System ◽  
Mutant Strain ◽  
Student Outcomes ◽  
Scientific Community ◽  
Undergraduate Research ◽  
Wild Type ◽  
Research Experiences ◽  
Undergraduate Research Experiences ◽  
The Impact ◽  
Insight Into

Course-based undergraduate research experiences (CUREs) have been shown to lead to multiple student benefits, but much is unknown about how CUREs lead to specific student outcomes. In this study, we examined the extent to which students making “broadly relevant novel discoveries” impacted student project ownership by comparing the experiences of students in a CURE and a traditional lab course. The CURE and traditional lab were similar in most aspects; students were exposed to an identical curriculum taught by the same instructor. However, there was one major difference between the two types of courses: the type of data that the students produced. Students in the traditional lab characterized the immune system of wild-type mice, thereby confirming results already known to the scientific community, while students in the CURE characterized the immune system of a mutant strain of mice, which produced broadly relevant novel discoveries. Compared with traditional lab students, CURE students reported higher cognitive and emotional ownership over their projects. Students’ perceptions of collaboration and making broadly relevant novel discoveries were significantly and positively related to their cognitive and emotional ownership. This work provides insight into the importance of integrating opportunities for broadly relevant novel discoveries in lab courses.

scholarly journals Epigenetic Control of Chromosome Breakage at the 5′ End of Paramecium tetraurelia Gene A

2004 ◽  
Vol 3 (5) ◽  
pp. 1136-1146 ◽  
Author(s):  
Laurence Amar ◽  
Karine Dubrana
Keyword(s):  
Mutant Strain ◽  
Type Strain ◽  
Wild Type Strain ◽  
Chromosome Breakage ◽  
Paramecium Tetraurelia ◽  
Wild Type ◽  
Dna Rearrangements ◽  
Rna Molecules ◽  
Extensive Analysis ◽  
Insight Into

ABSTRACT Macronuclei and micronuclei of ciliates have related genomes, with macronuclei developing from zygotic micronuclei through programmed DNA rearrangements. While Paramecium tetraurelia wild-type strain 51 and mutant strain d48 have the same micronuclear genome, qualitative differences between their macronuclear genomes have been described, demonstrating that programmed DNA rearrangements could be epigenetically controlled in ciliates. Macronuclear chromosomes end downstream of gene A (A51 Mac ends) and at the 5′ end of gene A (Ad48 Mac ends) in strains 51 and d48, respectively. To gain further insight into the process of chromosome end formation, we performed an extensive analysis of locus A rearrangement in strains d48 and 51, in strain d12, which harbors a gene A deletion, and in interstrain cross progeny. We show that (i) allele Ad12 harbors a deletion of >16 kb, (ii) A51 Mac ends distribute over four rather than three DNA regions, (iii) strains d48 and 51 display only quantitative differences (rare Ad48 and A51 Mac ends do form in strains 51 and d48, respectively), (iv) the level of A51 Mac ends is severalfold enhanced in d12- and d48-derived progeny, and (v) this level inversely correlates with the level of Ad48 Mac ends in the d48 parent. Together, these data lead to a model in which the formation of Ad48 Mac ends is epigenetically controlled by a d48 factor(s). We propose that the d48 factor(s) may be derived from RNA molecules transcribed from the Ad48 Mac ends and encompassing the truncated A gene and telomeric repeats.

Genetic analysis of developmental mechanisms in hydra. XIX. Stimulation of regeneration by injury in the regeneration-deficient mutant strain, reg-16

Development ◽  
1989 ◽  
Vol 105 (3) ◽  
pp. 521-528
Author(s):  
E. Kobatake ◽  
T. Sugiyama
Keyword(s):  
Mutant Strain ◽  
Deficient Mutant ◽  
Wild Type ◽  
Regenerative Capacity ◽  
Activation Level ◽  
Original Number ◽  
Morphogenetic Potential ◽  
Hydra Magnipapillata ◽  
Strain Mechanisms ◽  
Stimulation Of

A mutant strain of Hydra magnipapillata, reg-16, has a very low regenerative capacity. After head removal, it usually restores 10–20% of the original number of tentacles in 7 days. A procedure was found to markedly improve tentacle regeneration in this strain. The closed wound located at the apical regenerating tip of the decapitated polyp was gently reopened using a pair of forceps. Reg-16 polyps treated in this way at 24 and 48 h after head removal restored nearly all of the original number of tentacles in 7 days. A lateral tissue transplantation procedure was employed to examine the effect of wound reopening on the morphogenetic potential of decapitated reg-16 polyps. Wound reopening produced a significant increase in head activation level without producing a preceding decrease in head inhibition level. This and other observations suggest that the coupled activation-inhibition changes that normally occur after head removal from the wild-type hydra do not occur in this strain. Mechanisms responsible for the wound reopening effect and the absence of activation-inhibition coupling in the mutant strain reg-16 are discussed.

scholarly journals Role of Porphyromonas gingivalis FeoB2 in Metal Uptake and Oxidative Stress Protection

2006 ◽  
Vol 74 (7) ◽  
pp. 4214-4223 ◽  
Author(s):  
Jia He ◽  
Hiroshi Miyazaki ◽  
Cecilia Anaya ◽  
Fan Yu ◽  
W. Andrew Yeudall ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Mutant Strain ◽  
Parental Strain ◽  
Atmospheric Oxygen ◽  
Host Cells ◽  
Deficient Mutant ◽  
Wild Type ◽  
Stress Protection ◽  
Oxidative Stress Protection

ABSTRACT Porphyromonas gingivalis, a gram-negative anaerobic bacterium, is a recognized periodontopathogen. It exhibits a high degree of aerotolerance and is able to survive in host cells, indicating that efficient oxidative stress protection mechanisms must be present in this organism. Manganese homeostasis plays a major role in oxidative stress protection in a variety of organisms; however, the transport and role of this metal in P. gingivalis is not well understood. Analysis of the genome of P. gingivalis W83 revealed the presence of two genes encoding homologs of a ferrous iron transport protein, FeoB1 and FeoB2. FeoB2 has been implicated in manganese accumulation in P. gingivalis. We sought to determine the role of the FeoB2 protein in metal transport as well as its contribution to resistance to oxygen radicals. Quantitative reverse transcriptase PCR analyses demonstrated that expression of feoB2 is induced in the presence of oxygen. The role of FeoB2 was investigated using an isogenic mutant strain deficient in the putative transporter. We characterized the FeoB2-mediated metal transport using 55Fe2+ and 54Mn2+. The FeoB2-deficient mutant had dramatically reduced rates of manganese uptake (0.028 pmol/min/107 bacteria) compared with the parental strain (0.33 pmol/min/107 bacteria) (after 20 min of uptake using 50 nM of 54Mn2+). The iron uptake rates, however, were higher in the mutant strain (0.75 pmol/min/107 bacteria) than in the wild type (0.39 pmol/min/107 bacteria). Interestingly, reduced survival rates were also noted for the mutant strain after exposure to H2O2 and to atmospheric oxygen compared to the parental strain cultured under the same conditions. In addition, in vitro infection of host cells with the wild type, the FeoB2-deficient mutant, and the same-site revertant revealed that the mutant had a significantly decreased capability for intracellular survival in the host cells compared to the wild-type strain. Our results demonstrate that feoB2 encodes a major manganese transporter required for protection of the bacterium from oxidative stress generated by atmospheric oxygen and H2O2. Furthermore, we show that FeoB2 and acquisition of manganese are required for intracellular survival of P. gingivalis in host cells.

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