Some properties of a mitochondrial endonuclease from yeast

1982 ◽  
Vol 60 (7) ◽  
pp. 757-762 ◽  
Author(s):  
R. Morosoli ◽  
C.V. Lusena
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Ionic Strength ◽  
Nuclear Dna ◽  
Double Helix ◽  
Endonuclease Activity ◽  
Mt Dna ◽  
Isolated Mitochondria ◽  
Modified Dna ◽  
Dna Strands

A mitochondrial endonuclease from Saccharomyces cerevisiae was previously shown to cut both strands of native DNA at opposite or nearby sites. The present studies demonstrate that the endonuclease activity is dependent on the strength of the hydrogen bonds between the DNA strands; the activity was measured at different ionic strengths, with substrates of different base compositions and also with DNA in which the double helix has been locally destabilized by ultraviolet irradiation, by depurination, and by single-stranded nicks. The activity is 30% greater with mitochondrial DNA (mt-DNA) than with nuclear DNA. At 0.08 ionic strength, the relative activities with double-stranded polydeoxyribonucleotides are 2.4:1:0.6 for poly(dA)∙poly(dU): poly(dA)∙poly(dT): poly(dG)∙poly(dC). Increasing ionic strength decreases similarly me activity with poly(dA)∙poly(dU) and poly(dA)∙poly(dT), but has little effect with poly(dG)∙poly(dC). The greater activity with poly(dA)∙poly(dU) than with poly(dA)∙poly(dT) was confirmed with nick-translated mt-DNA and with DNA synthesized in isolated mitochondria using [3H]TTP and [3H]dUTP in both cases. The endonuclease cuts modified DNA preferentially in the thymine dinner regions, at the apurirtic sites, and at sites opposite to nicks. The possible involvement of this endonuclease in the degradation of mitochondrial DNA during "petite" induction is discussed.

Properties of Isolated Mitochondria of Agaricus Bisporus: Their Relationship to Dormancy and the Germination of Spores

1973 ◽  
Vol 31 ◽  
pp. 458-459
Author(s):  
K. S. McCarty ◽  
R. F. Weave ◽  
L. Kemper ◽  
F. S. Vogel
Keyword(s):  
Mitochondrial Dna ◽  
Ethidium Bromide ◽  
Microscopic Examination ◽  
Agaricus Bisporus ◽  
Osmotic Shock ◽  
Electron Microscopic Examination ◽  
Electron Microscopic ◽  
Isolated Mitochondria ◽  
Dna Strands ◽  
Cytoplasmic Ribosomes

During the prodromal stages of sporulation in the Basidiomycete, Agaricus bisporus, mitochondria accumulate in the basidial cells, zygotes, in the gill tissues prior to entry of these mitochondria, together with two haploid nuclei and cytoplasmic ribosomes, into the exospores. The mitochondria contain prominent loci of DNA [Fig. 1]. A modified Kleinschmidt spread technique1 has been used to evaluate the DNA strands from purified whole mitochondria released by osmotic shock, mitochondrial DNA purified on CsCl gradients [density = 1.698 gms/cc], and DNA purified on ethidium bromide CsCl gradients. The DNA appeared as linear strands up to 25 u in length and circular forms 2.2-5.2 u in circumference. In specimens prepared by osmotic shock, many strands of DNA are apparently attached to membrane fragments [Fig. 2]. When mitochondria were ruptured in hypotonic sucrose and then fixed in glutaraldehyde, the ribosomes were released for electron microscopic examination.

Comparison of the levels of the 21S mitochondrial rRNA in derepressed and glucose-repressed Saccharomyces cerevisiae

1983 ◽  
Vol 3 (11) ◽  
pp. 1949-1957
Author(s):  
R Kelly ◽  
S L Phillips
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Mitochondrial Function ◽  
Nuclear Dna ◽  
Glucose Repression ◽  
Cellular Level ◽  
Rrna Gene ◽  
Mitochondrial Rna ◽  
Mitochondrial Rrna ◽  
Cdna Preparation

A cDNA preparation, synthesized by using Saccharomyces cerevisiae mitochondrial RNA as template and oligodeoxythymidylic acid as primer, was found to specifically hybridize to the mitochondrial 21S rRNA by the following criteria: (i) it hybridizes only to the 21S RNA species in mitochondrial RNA and not to RNA from a [rho0] mutant, and (ii) it hybridizes to fragments in restriction digests of mitochondrial DNA that contain the 21S rRNA gene but not to nuclear DNA. This cDNA was used as a probe to demonstrate that a 2.6-fold decrease in the cellular level of the mitochondrial large rRNA is associated with glucose repression of mitochondrial function in S. cerevisiae. A corresponding decrease in the level of mitochondrial DNA was not observed.

Mitochondrial DNA synthesis in mouse L cells temperature sensitive in nuclear DNA replication

1977 ◽  
Vol 55 (5) ◽  
pp. 543-547 ◽  
Author(s):  
Rose Sheinin ◽  
Pamela Darragh ◽  
Margaret Dubsky
Keyword(s):  
Mitochondrial Dna ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Nuclear Dna ◽  
Temperature Sensitive ◽  
Chromosomal Dna ◽  
Mt Dna ◽  
L Cells ◽  
Mitochondrial Dna Synthesis

Temperature-sensitive (ts) A1S9 mouse L cells continue to synthesize double-stranded covalently closed mitochondrial (mt) DNA at a temperature (38.5 °C) which is nonpermissive for chromosomal DNA replication. The amount of mt DNA made appears to be quantitatively linked to nuclear DNA synthesis. Nuclear DNA replication proceeds normally for 6–8 h after the cells are shifted to 38.5 °C, and then declines to reach a minimum at 20–24 h. The level of mt DNA synthesis remains high during this period and decreases once the ts lesion has been established.

scholarly journals Stable transplantation of human mitochondrial DNA by high-throughput, pressurized isolated mitochondrial delivery

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Alexander J Sercel ◽  
Alexander N Patananan ◽  
Tianxing Man ◽  
Ting-Hsiang Wu ◽  
Amy K Yu ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Oxidative Phosphorylation ◽  
Mammalian Cells ◽  
Nuclear Dna ◽  
Cell Fates ◽  
Human Mitochondrial Dna ◽  
Isolated Mitochondria ◽  
Mitochondrial Transfer ◽  
Communication And Coordination ◽  
Transfer Device

Generating mammalian cells with specific mitochondrial DNA (mtDNA)–nuclear DNA (nDNA) combinations is desirable but difficult to achieve and would be enabling for studies of mitochondrial-nuclear communication and coordination in controlling cell fates and functions. We developed ‘MitoPunch’, a pressure-driven mitochondrial transfer device, to deliver isolated mitochondria into numerous target mammalian cells simultaneously. MitoPunch and MitoCeption, a previously described force-based mitochondrial transfer approach, both yield stable isolated mitochondrial recipient (SIMR) cells that permanently retain exogenous mtDNA, whereas coincubation of mitochondria with cells does not yield SIMR cells. Although a typical MitoPunch or MitoCeption delivery results in dozens of immortalized SIMR clones with restored oxidative phosphorylation, only MitoPunch can produce replication-limited, non-immortal human SIMR clones. The MitoPunch device is versatile, inexpensive to assemble, and easy to use for engineering mtDNA–nDNA combinations to enable fundamental studies and potential translational applications.

scholarly journals Comparison of the levels of the 21S mitochondrial rRNA in derepressed and glucose-repressed Saccharomyces cerevisiae.

1983 ◽  
Vol 3 (11) ◽  
pp. 1949-1957 ◽  
Author(s):  
R Kelly ◽  
S L Phillips
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Mitochondrial Function ◽  
Nuclear Dna ◽  
Glucose Repression ◽  
Cellular Level ◽  
Rrna Gene ◽  
Mitochondrial Rna ◽  
Mitochondrial Rrna ◽  
Cdna Preparation

A cDNA preparation, synthesized by using Saccharomyces cerevisiae mitochondrial RNA as template and oligodeoxythymidylic acid as primer, was found to specifically hybridize to the mitochondrial 21S rRNA by the following criteria: (i) it hybridizes only to the 21S RNA species in mitochondrial RNA and not to RNA from a [rho0] mutant, and (ii) it hybridizes to fragments in restriction digests of mitochondrial DNA that contain the 21S rRNA gene but not to nuclear DNA. This cDNA was used as a probe to demonstrate that a 2.6-fold decrease in the cellular level of the mitochondrial large rRNA is associated with glucose repression of mitochondrial function in S. cerevisiae. A corresponding decrease in the level of mitochondrial DNA was not observed.

scholarly journals Changes in circulating cell-free nuclear DNA and mitochondrial DNA of patients with adolescent idiopathic scoliosis

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Jiong Li ◽  
Longjie Wang ◽  
Guanteng Yang ◽  
Yunjia Wang ◽  
Chaofeng Guo ◽  
...  
Keyword(s):  
Mitochondrial Dna ◽  
Adolescent Idiopathic Scoliosis ◽  
Idiopathic Scoliosis ◽  
Sensitivity And Specificity ◽  
Dna Analysis ◽  
Nuclear Dna ◽  
Predictive Accuracy ◽  
Lower Sensitivity ◽  
Mt Dna ◽  
Potential Biomarker

Abstract Background Adolescent idiopathic scoliosis (AIS) which characterized by complex three-dimensional deformity of spine has been difficult to cure because of the unknown etiopathology and uncertainty of progression. Nowadays, circulating cell-free (ccf) DNA was found to be a potential biomarker for several benign and malignant diseases. However, whether ccf DNA can be a biomarker for AIS has not been reported yet. In this study, we investigate the circulating cell-free nuclear DNA (ccf n-DNA) and mitochondrial DNA (ccf mt-DNA) concentrations in the plasma of patients with AIS and controls (CT), and the changed plasma ccf n-DNA and ccf mt-DNA levels and their association with clinical parameters were assessed. Methods The plasma of peripheral blood from 69 AIS patients and 21 age-matched CT was collected for ccf DNA analysis. Quantitative PCR was used to detect ccf n-DNA and ccf mt-DNA levels, and correlation analyses between the ccf n-DNA and ccf mt-DNA levels and clinical characteristics were conducted. Receiver operator curves (ROC) were used to analyze the sensitivity and specificity of ccf n-DNA and ccf mt-DNA levels to different characteristics. Results The plasma ccf n-DNA levels of both GAPDH and ACTB were significantly decreased in AIS patients compared with those in controls, while the plasma ccf mt-DNA levels did not changed. According to sex-related analyses, the ccf n-DNA levels in male CT-M was higher than that in female CT and male AIS, but the ccf n-DNA levels in female AIS was not significantly changed when compared with male AIS or female CT. However, the concentration of ccf mt-DNA in female AIS increased significantly when compared with male AIS. Surprisingly, Lenke type-related analyses suggested that Lenke type 1 patients had lower ccf n-DNA levels, whereas Lenke type 5 patients had higher ccf mt-DNA levels compared with those of controls. However, a lower sensitivity and specificity of AIS predicted by ccf n-DNA or ccf mt-DNA levels was observed, whether in total, by sex, or by Lenke type. Conclusion Although with no/little predictive accuracy of AIS/progressed AIS by ccf DNA levels, significantly changed plasma ccf DNA levels were observed in AIS patients compared with those in controls.

scholarly journals Mitochondrial DNA duplication, recombination, and introgression during interspecific hybridization

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Silvia Bágeľová Poláková ◽  
Žaneta Lichtner ◽  
Tomáš Szemes ◽  
Martina Smolejová ◽  
Pavol Sulo
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Interspecific Hybridization ◽  
Mobile Elements ◽  
Variable Length ◽  
Mitochondrial Genomes ◽  
Free Standing ◽  
Saccharomyces Paradoxus ◽  
Mtdna Recombination

AbstractmtDNA recombination events in yeasts are known, but altered mitochondrial genomes were not completed. Therefore, we analyzed recombined mtDNAs in six Saccharomyces cerevisiae × Saccharomyces paradoxus hybrids in detail. Assembled molecules contain mostly segments with variable length introgressed to other mtDNA. All recombination sites are in the vicinity of the mobile elements, introns in cox1, cob genes and free standing ORF1, ORF4. The transplaced regions involve co-converted proximal exon regions. Thus, these selfish elements are beneficial to the host if the mother molecule is challenged with another molecule for transmission to the progeny. They trigger mtDNA recombination ensuring the transfer of adjacent regions, into the progeny of recombinant molecules. The recombination of the large segments may result in mitotically stable duplication of several genes.

Development of a mito-CRISPR system for generating mitochondrial DNA-deleted strain in Saccharomyces cerevisiae

2020 ◽  
Vol 85 (4) ◽  
pp. 895-901
Author(s):  
Takamitsu Amai ◽  
Tomoka Tsuji ◽  
Mitsuyoshi Ueda ◽  
Kouichi Kuroda
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Dna ◽  
Ethidium Bromide ◽  
Nuclear Genome ◽  
Target Sequence ◽  
Guide Rna ◽  
Crispr System ◽  
Mitochondrial Target ◽  
Gene Treatment ◽  
Mtdna Deletion

ABSTRACT Mitochondrial dysfunction can occur in a variety of ways, most often due to the deletion or mutation of mitochondrial DNA (mtDNA). The easy generation of yeasts with mtDNA deletion is attractive for analyzing the functions of the mtDNA gene. Treatment of yeasts with ethidium bromide is a well-known method for generating ρ° cells with complete deletion of mtDNA from Saccharomyces cerevisiae. However, the mutagenic effects of ethidium bromide on the nuclear genome cannot be excluded. In this study, we developed a “mito-CRISPR system” that specifically generates ρ° cells of yeasts. This system enabled the specific cleavage of mtDNA by introducing Cas9 fused with the mitochondrial target sequence at the N-terminus and guide RNA into mitochondria, resulting in the specific generation of ρ° cells in yeasts. The mito-CRISPR system provides a concise technology for deleting mtDNA in yeasts.

scholarly journals The Role of Mitochondria in Carcinogenesis

2021 ◽  
Vol 22 (10) ◽  
pp. 5100
Author(s):  
Paulina Kozakiewicz ◽  
Ludmiła Grzybowska-Szatkowska ◽  
Marzanna Ciesielka ◽  
Jolanta Rzymowska
Keyword(s):  
Prostate Cancer ◽  
Mitochondrial Dna ◽  
Nuclear Dna ◽  
Dna Polymorphisms ◽  
Gene Encoding ◽  
Dna Mutations ◽  
Nadh Ubiquinone Oxidoreductase ◽  
Gene Coi ◽  
The Impact

The mitochondria are essential for normal cell functioning. Changes in mitochondrial DNA (mtDNA) may affect the occurrence of some chronic diseases and cancer. This process is complex and not entirely understood. The assignment to a particular mitochondrial haplogroup may be a factor that either contributes to cancer development or reduces its likelihood. Mutations in mtDNA occurring via an increase in reactive oxygen species may favour the occurrence of further changes both in mitochondrial and nuclear DNA. Mitochondrial DNA mutations in postmitotic cells are not inherited, but may play a role both in initiation and progression of cancer. One of the first discovered polymorphisms associated with cancer was in the gene NADH-ubiquinone oxidoreductase chain 3 (mt-ND3) and it was typical of haplogroup N. In prostate cancer, these mutations and polymorphisms involve a gene encoding subunit I of respiratory complex IV cytochrome c oxidase subunit 1 gene (COI). At present, a growing number of studies also address the impact of mtDNA polymorphisms on prognosis in cancer patients. Some of the mitochondrial DNA polymorphisms occur in both chronic disease and cancer, for instance polymorphism G5913A characteristic of prostate cancer and hypertension.

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