Biology subjects

Khrapko, M.

Publications and source records attributed to Khrapko, M..

6 recordsLinked to original sources

bioRxiv2026

Positive germline selection of mtDNA mutations: evidence from the oocyte.

Purifying selection is the removal of detrimental mutations. Nuclear DNA mutations are purged by removing mutant individuals, embryos, or germline cells. Because mtDNA is present in numerous copies per cell, purifying selection of mtDNA mutations introduces an additional layer of complexity. The intracellular mutant fraction can change due to clonal expansion/reduction of mutant mtDNA molecules in germline cells. Different research groups have reported either negative (purifying), or positive (destructive) germline mtDNA selection. In this study, we use recently published high-fidelity data on mtDNA mutations in individual human oocytes from the Makova laboratory (Arbeithuber et al., 2025) to resolve this disagreement. To assess selection, we used the "selective expansion" metric St, which compares the average weighted rate of clonal expansion in a set of mutations tested for selection (e.g. non-synonymous mutations) to that of the set of synonymous mutations. We found that, on average, noncoding oocyte mutations in the control region expand faster than synonymous mutations, i.e., are under positive selection. Intriguingly, mutations in the coding region, the detrimental ones in particular, were also on average under positive selection. The prescence of average positive selection does not preclude purifying selection against individual mutations; it only indicates that positive selection dominates in aggregate mutational dynamics. Positive selection impacts the dynamics of de novo germline mutations that arise in primordial germ cells or oocytes and have not yet been inherited by the next generation. Contrastly, mutations that are passed to subsequent generations are under average purifying selection. We suggest that initial positive selection begets subsequent purifying selection by increasing intracellular mutant fractions to levels at which their deleterious effects become phenotypically apparent and can be efficiently removed.

genetics↗

bioRxiv2024

A comprehensive map of transient protein-protein interactions and their structural features in the E. coli 1-carbon metabolism pathway

Enzymes in a pathway often form metabolons through weak protein-protein interactions (PPI) that localize and protect labile metabolites. Due to their transient nature, the structural architecture of these enzyme assemblies has largely remained elusive, limiting our abilities to re-engineer novel metabolic pathways. Here we delineate a complete PPI map of 1225 interactions in the E. coli 1-carbon metabolism pathway using bimolecular fluorescence complementation that can capture transient interactions in vivo and show strong intra- and inter-pathway clusters within the folate and purine biosynthesis pathways. Scanning mutagenesis experiments along with AlphaFold predictions and meta-dynamics simulations reveal that most proteins use conserved "dedicated" interfaces distant from their active sites to interact with multiple partners. Diffusion-reaction simulations with shared interaction surfaces and realistic PPI networks reveal a dramatic speedup in metabolic pathway fluxes. Overall, this study sheds light on the fundamental features of metabolon biophysics and structural aspects of transient binary complexes. Significance statementEnzymes from the same metabolic pathway often form dynamic assemblies called metabolons, which channel metabolites as well as protect labile intermediates. Yet very little is known about their structural features or what makes these interactions transient. Paucity of such information has particularly affected our ability to engineer novel metabolic pathways, construct multi-scale mathematical models of cells, etc. We address this by obtaining a comprehensive map for 1225 interaction pairs in the essential 1-carbon metabolism pathway of E. coli. Using both high-throughput experiments and computation, we uncover that metabolon proteins tend to use a conserved dedicated interface to interact with their partners. These results shed light on structural and energetic aspects of PPI in metabolons at near atomic level of resolution.

biophysics↗

bioRxiv2023

A novel, wave-shaped profile of germline selection of pathogenic mtDNA mutations is discovered by bypassing a classical statistical bias.

The shift of the level of disease-causing mtDNA mutations (heteroplasmy) from mother to child is typically negatively correlated with the mothers heteroplasmy (Hm). In other words, mothers with low Hm tend to have children with a higher mutation level (Hch) than their own. In contrast, mothers with high Hm typically see a decrease in heteroplasmy in their children. This trend has been commonly interpreted as a result of a descending germline selection profile, i.e., positive selection at low Hm, gradually turning negative at high Hm. Here we demonstrate, however, that the negative correlation is mostly driven by RTM, or Regression To the Mean, a classical statistical bias. We further show that RTM can be nullified by using the average between the mothers and childs heteroplasmy, as a new variable, instead of the commonly used mothers heteroplasmy in blood. Additionally, we demonstrate that mother/child average is a better approximation of the actual germline heteroplasmy. Moreover, the elimination of RTM revealed a previously hidden wave-shaped HS-profile (positive mother-to-child shift at intermediate average mother-child heteroplasmy, decreasing towards high and low average heteroplasmy). In confirmation of this finding, we show that simulations that involve both wave-shaped HS-profile and RTM, reproduce the observed patterns of inheritance of mtDNA mutations in unprecedented detail. From the health care perspective, the uncovering of the wave-shaped HS-profile (and the removal of the RTM bias) are crucial for families affected by mtDNA disease. From the fundamental perspective, the wave- shaped profile offers a novel understanding of the dynamics of mtDNA in the germline and a novel potential mechanism that prevents the spread of detrimental mtDNA mutations in the population. SignificanceFrom the clinical perspective, the existence of wave-shaped selection may improve predictions and decisions for families affected by mtDNA diseases. From the fundamental perspective, it provides insight into the dynamics of general mtDNA mutations in the germline and in the population, as long as they follow wave-shaped selection profile. In Fig. 1, blue and red arrows represent the direction of expected changes of the heteroplasmy in a lineage with time/generations. With wave- shaped selection (Fig. 1B), a great majority of nascent low- fraction mutations are expected to converge back to zero and vanish. However, due to random intracellular genetic drift, some mutations will, occasionally, expand and enter the range of positive selection. Then they will be expanded by the selection to higher, detrimental levels, and become prone to downstream removal via death of highly mutated germ cells or inability of highly sick individuals to continue their lineage. In this way, the wave-shaped selection may help to prevent the spread of detrimental mutations in the population and in the species. In contrast, if the descending selection profile (Fig. 1A) was in effect, the nascent low heteroplasmy detrimental mutations would have been pushed to intermediate heteroplasmy levels where they will stay longer in hidden disease carriers enabling effective spread of mutation in the population. O_FIG O_LINKSMALLFIG WIDTH=192 HEIGHT=200 SRC="FIGDIR/small/568140v2_fig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@1142cc0org.highwire.dtl.DTLVardef@186002aorg.highwire.dtl.DTLVardef@74d176org.highwire.dtl.DTLVardef@163ca6c_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1.C_FLOATNO (Graphic Summary) Pathogenic mtDNA mutations that cause a host of devastating inherited diseases are usually thought to follow an intriguing inheritance trend: Mothers with low levels of mutation (called mother heteroplasmy, Hm) tend to bear children with higher child heteroplasmy (Hch) then their own which constitutes positive Heteroplasmy Shift (HS=Hch-Hm). In contrast, mothers with high heteroplasmy Hm bear children with lower heteroplasmy Hc (negative HS). C_FIG

genomics↗

bioRxiv2022

Reanalysis of mtDNA mutations of human primordial germ cells (PGCs) reveals significant contamination with NUMTs, and challenges predominantly purifying selection in late PGCs

The resilience of the mitochondrial genome (mtDNA) to a high mutational pressure depends, in part, on negative purifying selection in the germline. A paradigm in the field has been that such selection, at least in part, takes place in primordial germ cells (PGCs). Specifically, Floros et al. (Nature Cell Biology 20: 144-51) reported an increase in the synonymity of mtDNA mutations (a sign of purifying selection) between pooled early-stage and late-stage PGCs. We re-analyzed Floros et al. pooled PGC data and noticed that their mutational dataset was significantly contaminated with single nucleotide variants (SNVs) derived from a nuclear sequence of mtDNA origin (NUMT) located on chromosome 5. Contamination was caused by co-amplification of the NUMT sequence by cross-specific PCR primers. Importantly, when we removed NUMT-derived SNVs, the evidence of purifying selection was abolished. In addition to pooled PGCs, Floros et al. reported the analysis of single late-stage PGCs, which were amplified with different sets of PCR primers that cannot amplify the NUMT sequence. Accordingly, we found no NUMT-derived SNVs among single PGCs mutations. Interestingly, single PGC mutations show a decrease of synonymity with increased intracellular mutant fraction. This pattern is incompatible with predominantly negative selection. This suggests that germline selection of mtDNA mutations is a complex phenomenon and that the part of this process that takes place in PGCs may be predominantly positive. However counterintuitive, positive germline selection of detrimental mtDNA mutations has been reported previously and potentially may be evolutionarily advantageous.

genomics↗

bioRxiv2021

Bi-phasic dynamics of the mitochondrial DNA mutation m.3243A>G in blood: An unbiased, mutation level-dependent model implies positive selection in the germline.

The A-to-G point mutation at position 3243 in the human mitochondrial genome (m.3243A>G) is the most common pathogenic mtDNA variant responsible for disease in humans. It is widely accepted that m.3243A>G levels decrease in blood with age, and an age correction representing [~]2% annual decline is often applied to account for this change in mutation level. Here we report that recent data indicate the dynamics of m.3243A>G are more complex and depend on the mutation level in blood in a bi-phasic way. Consequently, the traditional 2% correction, which is adequate on average, creates opposite predictive biases at high and low mutation levels. Unbiased age correction is needed to circumvent these drawbacks of the standard model. We propose to eliminate both biases by using an approach where age correction depends on mutation level in a biphasic way to account for the dynamics of m.3243A>G in blood. The utility of this approach was further tested in estimating germline selection of m.3243A>G. The biphasic approach permitted us to uncover patterns consistent with the possibility of positive selection for m.3243A>G. Germline selection of m.3243A>G shows an arching profile by which selection is positive at intermediate mutant fractions and declines at high and low mutant fractions. We conclude that use of this biphasic approach will greatly improve the accuracy of modelling changes in mtDNA mutation frequencies in the germline and in somatic cells during aging.

genomics↗

bioRxiv2019

Quasi-Mendelian Paternal Inheritance of mitochondrial DNA: A notorious artifact, or anticipated mtDNA behavior?

A recent report by Luo et al (2018) in PNAS (DOI:10.1073/pnas.1810946115) presented evidence of biparental inheritance of mitochondrial DNA. The pattern of inheritance, however, resembled that of a nuclear gene. The authors explained this peculiarity with Mendelian segregation of a faulty gatekeeper gene that permits survival of paternal mtDNA in the oocyte. Three other groups (Vissing, 2019; Lutz-Bonengel and Parson, 2019; Salas et al, 2019), however, posited the observation was an artifact of inheritance of mtDNA nuclear pseudogenes (NUMTs), present in the fathers nuclear genome. We present justification that both interpretations are incorrect, but that the original authors did, in fact, observe biparental inheritance of mtDNA. Our alternative model assumes that because of initially low paternal mtDNA copy number these copies are randomly partitioned into nascent cell lineages. The paternal mtDNA haplotype must have a selective advantage, so seeded cells will tend to proceed to fixation of the paternal haplotype in the course of development. We use modeling to emulate the dynamics of paternal genomes and predict their mode of inheritance and distribution in somatic tissue. The resulting offspring is a mosaic of cells that are purely maternal or purely paternal - including in the germline. This mosaicism explains the quasi-Mendelian segregation of the paternal mDNA. Our model is based on known aspects of mtDNA biology and explains all of the experimental observations outlined in Luo et. al., including maternal inheritance of the grand-paternal mtDNA.

genetics↗