Cell-cycle and Age-Related Modulations in Mouse Chromosome Stiffness
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eLife assessment
This valuable paper describes the stiffness of meiotic chromosomes in both oocytes and spermatocytes. The authors identify differences in stiffness between meiosis I and II chromosomes, as well as an age-dependent increase in stiffness in meiosis I (and meiosis II) chromosomes, results that are highly significant for the field of chromosome biology. The mechanisms underlying age-dependent changes in chromosome stiffness remain unclear, and the evidence to suggest that changes in stiffness are independent of cohesin, which is known to deteriorate with age, is incomplete.
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Abstract
The intricate structure of chromosomes is complex, and many aspects of chromosome configuration/organization remain to be fully understood. Measuring chromosome stiffness can provide valuable insights into their structure. However, the nature of chromosome stiffness, whether static or dynamic, remains elusive. In this study, we analyzed chromosome stiffness in MI and MII oocytes. We revealed that MI oocytes had a ten-fold increase in stiffness compared to mitotic chromosomes, whereas chromosome stiffness in MII oocytes was relatively low chromosome. We then investigated the contribution of meiosis-specific cohesin complexes to chromosome stiffness in MI and MII oocytes. Surprisingly, the Young’s modulus of chromosomes from the three meiosis-specific cohesin mutants did not exhibit significant differences compared to the wild type, indicating that these proteins may not play a substantial role in determining chromosome stiffness. Additionally, our findings revealed an age-related increase in chromosome stiffness in MI oocytes. Age correlates with elevated DNA damage levels, so we investigated the impact of etoposide-induced DNA damage on chromosome stiffness, discovering a reduction in stiffness in response to such damage in MI oocytes. Overall, our study underscores the dynamic nature of chromosome stiffness, subject to changes influenced by the cell cycle and age.
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eLife assessment
This valuable paper describes the stiffness of meiotic chromosomes in both oocytes and spermatocytes. The authors identify differences in stiffness between meiosis I and II chromosomes, as well as an age-dependent increase in stiffness in meiosis I (and meiosis II) chromosomes, results that are highly significant for the field of chromosome biology. The mechanisms underlying age-dependent changes in chromosome stiffness remain unclear, and the evidence to suggest that changes in stiffness are independent of cohesin, which is known to deteriorate with age, is incomplete.
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Reviewer #1 (Public Review):
Summary:
By using the biophysical chromosome stretching, the authors measured the stiffness of chromosomes of mouse oocytes in meiosis I (MI) and meiosis II (MII). This study was the follow-up of previous studies in spermatocytes (and oocytes) by the authors (Biggs et al. Commun. Biol. 2020: Hornick et al. J. Assist. Rep. and Genet. 2015). They showed that MI chromosomes are much stiffer (~10 fold) than mitotic chromosomes of mouse embryonic fibroblast (MEF) cells. MII chromosomes are also stiffer than the mitotic chromosomes. The authors also found that oocyte aging increases the stiffness of the chromosomes. Surprisingly, the stiffness of meiotic chromosomes is independent of meiotic chromosome components, Rec8, Stag3, and Rad21L. with aging.
Strengths:
This provides a new insight into the biophysical …
Reviewer #1 (Public Review):
Summary:
By using the biophysical chromosome stretching, the authors measured the stiffness of chromosomes of mouse oocytes in meiosis I (MI) and meiosis II (MII). This study was the follow-up of previous studies in spermatocytes (and oocytes) by the authors (Biggs et al. Commun. Biol. 2020: Hornick et al. J. Assist. Rep. and Genet. 2015). They showed that MI chromosomes are much stiffer (~10 fold) than mitotic chromosomes of mouse embryonic fibroblast (MEF) cells. MII chromosomes are also stiffer than the mitotic chromosomes. The authors also found that oocyte aging increases the stiffness of the chromosomes. Surprisingly, the stiffness of meiotic chromosomes is independent of meiotic chromosome components, Rec8, Stag3, and Rad21L. with aging.
Strengths:
This provides a new insight into the biophysical property of meiotic chromosomes, that is chromosome stiffness. The stiffness of chromosomes in meiosis prophase I is ~10-fold higher than that of mitotic chromosomes, which is independent of meiotic cohesin. The increased stiffness during oocyte aging is a novel finding.
Weaknesses:
A major weakness of this paper is that it does not provide any molecular mechanism underlying the difference between MI and MII chromosomes (and/or prophase I and mitotic chromosomes).
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Reviewer #2 (Public Review):
This paper reports investigations of chromosome stiffness in oocytes and spermatocytes. The paper shows that prophase I spermatocytes and MI/MII oocytes yield high Young Modulus values in the assay the authors applied. Deficiency in each one of three meiosis-specific cohesins they claim did not affect this result and increased stiffness was seen in aged oocytes but not in oocytes treated with the DNA-damaging agent etoposide.
The paper reports some interesting observations which are in line with a report by the same authors of 2020 where increased stiffness of spermatocyte chromosomes was already shown. In that sense, the current manuscript is an extension of that previous paper, and thus novelty is somewhat limited. The paper is also largely descriptive as it does neither propose a mechanism nor report …
Reviewer #2 (Public Review):
This paper reports investigations of chromosome stiffness in oocytes and spermatocytes. The paper shows that prophase I spermatocytes and MI/MII oocytes yield high Young Modulus values in the assay the authors applied. Deficiency in each one of three meiosis-specific cohesins they claim did not affect this result and increased stiffness was seen in aged oocytes but not in oocytes treated with the DNA-damaging agent etoposide.
The paper reports some interesting observations which are in line with a report by the same authors of 2020 where increased stiffness of spermatocyte chromosomes was already shown. In that sense, the current manuscript is an extension of that previous paper, and thus novelty is somewhat limited. The paper is also largely descriptive as it does neither propose a mechanism nor report factors that determine the chromosomal stiffness.
There are several points that need to be considered.
(1) Limitations of the study and the conclusions are not discussed in the "Discussion" section and that is a significant gap. Even more so as the authors rely on just one experimental system for all their data - there is no independent verification - and that in vitro system may be prone to artefacts.
(2) It is somewhat unfortunate that they jump between oocytes and spermatocytes to address the cohesin question. Prophase I (pachytene) spermatocytes chromosomes are not directly comparable to MI or MII oocyte chromosomes. In fact, the authors report Young Modulus values of 3700 for MI oocytes and only 2700 for spermatocyte prophase chromosomes, illustrating this difference. Why not use oocyte-specific cohesin deficiencies?
(3) It remains unclear whether the treatment of oocytes with the detergent TritonX-100 affects the spindle and thus the chromosomes isolated directly from the Triton-lysed oocytes. In fact, it is rather likely that the detergent affects chromatin-associated proteins and thus structural features of the chromosomes.
(4) Why did the authors use mouse strains of different genetic backgrounds, CD-1, and C57BL/6? That makes comparison difficult. Breeding of heterozygous cohesin mutants will yield the ideal controls, i.e. littermates.
(5) How did the authors capture chromosome axes from STAG3-deficienct spermatocytes which feature very few if any axes? How representative are those chromosomes that could be captured?
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Reviewer #3 (Public Review):
Summary:
Understanding the mechanical properties of chromosomes remains an important issue in cell biology. Measuring chromosome stiffness can provide valuable insights into chromosome organization and function. Using a sophisticated micromanipulation system, Liu et al. analyzed chromosome stiffness in MI and MII oocytes. The authors found that chromosomes in MI oocytes were ten-fold stiffer than mitotic ones. The stiffness of chromosomes in MI mouse oocytes was significantly higher than that in MII oocytes. Furthermore, the knockout of the meiosis-specific cohesin component (Rec8, Stag3, Rad21l) did not affect meiotic chromosome stiffness. Interestingly, the authors showed that chromosomes from old MI oocytes had higher stiffness than those from young MI oocytes. The authors claimed this effect was not due …
Reviewer #3 (Public Review):
Summary:
Understanding the mechanical properties of chromosomes remains an important issue in cell biology. Measuring chromosome stiffness can provide valuable insights into chromosome organization and function. Using a sophisticated micromanipulation system, Liu et al. analyzed chromosome stiffness in MI and MII oocytes. The authors found that chromosomes in MI oocytes were ten-fold stiffer than mitotic ones. The stiffness of chromosomes in MI mouse oocytes was significantly higher than that in MII oocytes. Furthermore, the knockout of the meiosis-specific cohesin component (Rec8, Stag3, Rad21l) did not affect meiotic chromosome stiffness. Interestingly, the authors showed that chromosomes from old MI oocytes had higher stiffness than those from young MI oocytes. The authors claimed this effect was not due to the accumulated DNA damage during the aging process because induced DNA damage reduced chromosome stiffness in oocytes.
Strengths:
The technique used (isolating the chromosomes in meiosis and measuring their stiffness) is the authors' specialty. The results are intriguing and informative to the chromatin/chromosome and other related fields.
Weaknesses:
(1) How intact the measured chromosomes were is unclear.
(2) Some control data needs to be included.
(3) The paper was not well-written, particularly the Introduction section.
(4) How intact were the measured chromosomes? Although the structural preservation of the chromosomes is essential for this kind of measurement, the meiotic chromosomes were isolated in PBS with Triton X-100 and measured at room temperature. It is known that chromosomes are very sensitive to cation concentrations and macromolecular crowding in the environment (PMID: 29358072, 22540018, 37986866). It would be better to discuss this point.
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