AP19676334 Study of the DNA strands inheritance in human mitochondrial DNA
Project goal: The aim of the present project is to investigate the DNA strand equivalence of mitochondrial genome of human and yeast cells by applying the logic of Meselson-Stahl experiment.
Project description: Mitochondria are membrane-bound cell organelles that contain their own genomic DNA, referred to as mitochondrial (mt) DNA with unique replication, transcription and translational machinery. Human cell contains several thousand copies of mtDNA, which is organised as a small closed circular DNA duplex of 16,569 bp. An intriguing peculiarity of the vertebrate mitochondrial genome is an unusual misbalance of nucleotide composition on the two strands of the mtDNA, which results in the separation into a heavy (H) G+T-rich and a light (L) C+A-rich DNA strand upon ultracentrifugation in an alkaline cesium chloride gradient. DNA replication in human mitochondria was investigated for several decades; nevertheless, at present, its mechanism and possible relationship to the skewed nucleotide composition remain subject of debate. Previously, we proposed a model of asymmetric DNA strand inheritance or non-equivalence of DNA strands, which provides a simple, heuristic explanation to the highly biased pattern of mutations in vertebrate mtDNA. Our model is consistent with the strand-asynchronous models of mtDNA replication described previously, but contains several new important features, which apparently have not been discussed before. We propose that the H-strand of mtDNA is used as a template for replication more often than the L-strand. Consequently, H-strand breeds the majority of mtDNA progeny, whereas the L-strand of mtDNA is served mainly for transcription and undergoes only one cycle of replication thus leaving few progeny behind. This model provides insight into the origin of skewed nucleotide composition of human mitochondrial genome and to the strand bias in the somatic mutations observed in the mtDNA from aging brain and cancer cells.
Project facilitators: PI: Sabira TAIPAKOVA, Murat SAPARBAEV, Alexander ISHCHENKO, Co-PI: Bakhyt MATKARIMOV, Zhanat KOSHENOV, Eldar BAIKEN, Diana MANAPKYZY, Botagoz DJOLDYBAEVA, Aizhan ALIKUL
Realisation period: 2023-2025
Expected results: Science-wise, the major impacts through the research activities of this project are:
(A) Identification of the preferential bias for the replication of H-strand of mtDNA in human cells, which might explain the mutation pattern and unusual nucleotide composition of vertebrate mtDNA;
(B) Demonstration of the DNA strand equivalence in nucDNA of human and yeast cells, which might explain intra-strand DNA symmetry and patterns of mutations in cellular organisms;
(C) Study of mtDNA replication in post-mitotic cells and in cells exposed to DNA damage will shed light on the mechanisms of regulation of DNA strand equivalence and inheritance.
As far as technological impact of this project is concerned, we will develop and validate the method to measure DNA strand equivalence in mtDNA in living cells. This new tool could be used as a diagnostic marker for mitochondrial diseases. The socioeconomic benefits of the present project would be to define a novel therapeutic paradigm. The modulation of mtDNA replication may have different impacts: increasing the preference for replication of H-strand may increase purifying selection and reduce heteroplasmy; whereas more balanced replication of H- and L-strands may increase the rate of deleterious mtDNA mutations and this could be used in anti-cancer therapy. The knowledge produced in this project on the mechanisms of mtDNA strand inheritance will help to understand patterns of somatic mutations in nuclear and mtDNA, this in turn will help to identify novel diagnostic and therapeutic strategies to combat age-related and mitochondrial diseases.
According to results of the project at least 3 (three) articles and (or) reviews in peer-reviewed scientific publications in the scientific direction of the project, indexed in the Science Citation Index Expanded and included in the 1st (first), 2nd (second) and (or) 3rd (third) quartile by impact factor in the Web of Science database and (or) having a CiteScore percentile in Scopus database of at least 50 (fifty); or at least 2 (two) articles and (or) reviews in peer-reviewed scientific publications indexed in the Science Citation Index Expanded and included in the 1st (first) and (or) 2nd (second) quartile by impact factor in the Web of Science database and (or) having a CiteScore percentile in the Scopus database of at least 65 (sixty-five); or at least 1 (one) article or review in a peer-reviewed scientific publication indexed in the Science Citation Index Expanded and included in the 1st (first) quartile in the Web of Science database or having a CiteScore percentile in the Scopus database of at least 95 (ninety-five).
In addition, at least 1 (one) article or review in a peer-reviewed foreign or domestic publication recommended by the KOKSNVO.
Methodology: Human and yeast cell culturing, cellular DNA isolation, highly sensitive LC-MS. Exonuclease III mediated conversion of dsDNA to ssDNA, Denaturing gel electrophoresis, Southern blot, affinity chromatography. Human and yeast cell culture, cellular DNA isolation, DNA strand separation, LC-MS and/or LC-MSn. Cells treatment with DNA damaging agents, mtDNA purification, DNA strands separation, HPLC-MSn analysis of nucleosides from ssDNA.
Contacts: sabira.taipakova@gmail.com
Project goal: The aim of the present project is to investigate the DNA strand equivalence of mitochondrial genome of human and yeast cells by applying the logic of Meselson-Stahl experiment.
Project description: Mitochondria are membrane-bound cell organelles that contain their own genomic DNA, referred to as mitochondrial (mt) DNA with unique replication, transcription and translational machinery. Human cell contains several thousand copies of mtDNA, which is organised as a small closed circular DNA duplex of 16,569 bp. An intriguing peculiarity of the vertebrate mitochondrial genome is an unusual misbalance of nucleotide composition on the two strands of the mtDNA, which results in the separation into a heavy (H) G+T-rich and a light (L) C+A-rich DNA strand upon ultracentrifugation in an alkaline cesium chloride gradient. DNA replication in human mitochondria was investigated for several decades; nevertheless, at present, its mechanism and possible relationship to the skewed nucleotide composition remain subject of debate. Previously, we proposed a model of asymmetric DNA strand inheritance or non-equivalence of DNA strands, which provides a simple, heuristic explanation to the highly biased pattern of mutations in vertebrate mtDNA. Our model is consistent with the strand-asynchronous models of mtDNA replication described previously, but contains several new important features, which apparently have not been discussed before. We propose that the H-strand of mtDNA is used as a template for replication more often than the L-strand. Consequently, H-strand breeds the majority of mtDNA progeny, whereas the L-strand of mtDNA is served mainly for transcription and undergoes only one cycle of replication thus leaving few progeny behind. This model provides insight into the origin of skewed nucleotide composition of human mitochondrial genome and to the strand bias in the somatic mutations observed in the mtDNA from aging brain and cancer cells.
Project facilitators: PI: Sabira TAIPAKOVA, Murat SAPARBAEV, Alexander ISHCHENKO, Co-PI: Bakhyt MATKARIMOV, Zhanat KOSHENOV, Eldar BAIKEN, Diana MANAPKYZY, Botagoz DJOLDYBAEVA, Aizhan ALIKUL
Realisation period: 2023-2025
Expected results: Science-wise, the major impacts through the research activities of this project are:
(A) Identification of the preferential bias for the replication of H-strand of mtDNA in human cells, which might explain the mutation pattern and unusual nucleotide composition of vertebrate mtDNA;
(B) Demonstration of the DNA strand equivalence in nucDNA of human and yeast cells, which might explain intra-strand DNA symmetry and patterns of mutations in cellular organisms;
(C) Study of mtDNA replication in post-mitotic cells and in cells exposed to DNA damage will shed light on the mechanisms of regulation of DNA strand equivalence and inheritance.
As far as technological impact of this project is concerned, we will develop and validate the method to measure DNA strand equivalence in mtDNA in living cells. This new tool could be used as a diagnostic marker for mitochondrial diseases. The socioeconomic benefits of the present project would be to define a novel therapeutic paradigm. The modulation of mtDNA replication may have different impacts: increasing the preference for replication of H-strand may increase purifying selection and reduce heteroplasmy; whereas more balanced replication of H- and L-strands may increase the rate of deleterious mtDNA mutations and this could be used in anti-cancer therapy. The knowledge produced in this project on the mechanisms of mtDNA strand inheritance will help to understand patterns of somatic mutations in nuclear and mtDNA, this in turn will help to identify novel diagnostic and therapeutic strategies to combat age-related and mitochondrial diseases.
According to results of the project at least 3 (three) articles and (or) reviews in peer-reviewed scientific publications in the scientific direction of the project, indexed in the Science Citation Index Expanded and included in the 1st (first), 2nd (second) and (or) 3rd (third) quartile by impact factor in the Web of Science database and (or) having a CiteScore percentile in Scopus database of at least 50 (fifty); or at least 2 (two) articles and (or) reviews in peer-reviewed scientific publications indexed in the Science Citation Index Expanded and included in the 1st (first) and (or) 2nd (second) quartile by impact factor in the Web of Science database and (or) having a CiteScore percentile in the Scopus database of at least 65 (sixty-five); or at least 1 (one) article or review in a peer-reviewed scientific publication indexed in the Science Citation Index Expanded and included in the 1st (first) quartile in the Web of Science database or having a CiteScore percentile in the Scopus database of at least 95 (ninety-five).
In addition, at least 1 (one) article or review in a peer-reviewed foreign or domestic publication recommended by the KOKSNVO.
Methodology: Human and yeast cell culturing, cellular DNA isolation, highly sensitive LC-MS. Exonuclease III mediated conversion of dsDNA to ssDNA, Denaturing gel electrophoresis, Southern blot, affinity chromatography. Human and yeast cell culture, cellular DNA isolation, DNA strand separation, LC-MS and/or LC-MSn. Cells treatment with DNA damaging agents, mtDNA purification, DNA strands separation, HPLC-MSn analysis of nucleosides from ssDNA.
Contacts: sabira.taipakova@gmail.com