mTRIP: An Imaging Tool to Investigate Mitochondrial DNA Dynamics in Physiology and Disease at the Single-Cell Resolution

Atomic force microscopy (AFM) is a widely used imaging technique in material sciences. After becoming a standard surface-imaging tool, AFM has been proven to be useful in addressing several biological issues such as the characterization of cell organelles, quantification of DNA-protein interactions, cell adhesion forces, and electromechanical properties of living cells. AFM technique has undergone many successful improvements since its invention, including fluidic force microscopy (FluidFM), which combines conventional AFM with microchanneled cantilevers for local liquid dispensing. This technology permitted to overcome challenges linked to single-cell analyses. Indeed, FluidFM allows isolation and injection of single cells, force-controlled patch clamping of beating cardiac cells, serial weighting of micro-objects, and single-cell extraction for molecular analyses. This work aims to review the recent studies of AFM implementation in molecular and cellular biology.

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Heteroplasmy variability in individuals with biparentally inherited mitochondrial DNA

10.1101/2020.02.26.939405 ◽

2020 ◽

Author(s):

Jesse Slone ◽

Weiwei Zou ◽

Shiyu Luo ◽

Eric S Schmitt ◽

Stella Maris Chen ◽

...

Keyword(s):

Mitochondrial Dna ◽

Single Cell ◽

Strong Evidence ◽

Genetic Factors ◽

Single Cell Level ◽

Biparental Inheritance ◽

Cell Level ◽

Single Population ◽

Parent Of Origin ◽

Two Populations

ABSTRACTWith very few exceptions, mitochondrial DNA (mtDNA) in humans is transmitted exclusively from mothers to their offspring, suggesting the presence of a strong evolutionary pressure favoring the exclusion of paternal mtDNA. We have recently shown strong evidence of paternal mtDNA transmission. In these rare situations, males exhibiting biparental mtDNA appear to be limited to transmitting just one of the mtDNA species to their offspring, while females possessing biparental mtDNA populations consistently transmit both populations to their offspring at a very similar heteroplasmy level. The precise biological and genetic factors underlying this unusual transmission event remain unclear. Here, we have examined heteroplasmy levels in various tissues among individuals with biparental inheritance. Our results indicate that individuals with biparental mtDNA have remarkable inter-tissue variability in heteroplasmy level. At the single-cell level, paternal mtDNA heteroplasmy in sperm varies dramatically, and many sperm possess only one of the two mtDNA populations originally in question. These results show a fundamental, parent-of-origin difference in how mtDNA molecules transmit and propagate. This helps explain how a single population of mtDNAs are transmitted from a father possessing two populations of mtDNA molecules, suggesting that some mtDNA populations may be favored over others when transmitted from the father.

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Understanding mitochondrial DNA maintenance disorders at the single muscle fibre level

Nucleic Acids Research ◽

10.1093/nar/gkz472 ◽

2019 ◽

Vol 47 (14) ◽

pp. 7430-7443 ◽

Cited By ~ 5

Author(s):

Diana Lehmann ◽

Helen A L Tuppen ◽

Georgia E Campbell ◽

Charlotte L Alston ◽

Conor Lawless ◽

...

Keyword(s):

Mitochondrial Dna ◽

Single Cell ◽

Respiratory Chain ◽

Genetic Defect ◽

Clear Correlation ◽

Muscle Fibres ◽

Single Muscle Fibre ◽

Respiratory Chain Deficiency ◽

Mtdna Deletions ◽

Mtdna Deletion

Abstract Clonal expansion of mitochondrial DNA (mtDNA) deletions is an important pathological mechanism in adults with mtDNA maintenance disorders, leading to a mosaic mitochondrial respiratory chain deficiency in skeletal muscle. This study had two aims: (i) to determine if different Mendelian mtDNA maintenance disorders showed similar pattern of mtDNA deletions and respiratory chain deficiency and (ii) to investigate the correlation between the mitochondrial genetic defect and corresponding respiratory chain deficiency. We performed a quantitative analysis of respiratory chain deficiency, at a single cell level, in a cohort of patients with mutations in mtDNA maintenance genes. Using the same tissue section, we performed laser microdissection and single cell genetic analysis to investigate the relationship between mtDNA deletion characteristics and the respiratory chain deficiency. The pattern of respiratory chain deficiency is similar with different genetic defects. We demonstrate a clear correlation between the level of mtDNA deletion and extent of respiratory chain deficiency within a single cell. Long-range and single molecule PCR shows the presence of multiple mtDNA deletions in approximately one-third of all muscle fibres. We did not detect evidence of a replicative advantage for smaller mtDNA molecules in the majority of fibres, but further analysis is needed to provide conclusive evidence.

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