Review Reports - Nuclear Mechanics and Nuclear Mechanotransduction in Cancer Cell Migration and Invasion

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

A review manuscript titled, Nuclear mechanics and nuclear mechanotransduction in cancer cell migration and invasion’ by the author Claudia Tanja Mierke aimed to study the role of nuclear deformation during cancer dissemination. The manuscript started with an introduction covering mechanosensation, mechano-transduction and nuclear mechanics, followed by their mechanical insights into their roles in different event of biomechanism. Further, the author discussed about traditional characteristics of the nucleus are used as biomarkers for cancer, mechanical characteristics, nuclear bodies, nucleoplasm and chromatin, the nucleus functions, and related events. The author has described the knowledge gap, and concluded this kind of research. The manuscript also included sufficient Figures and presented, clearly. The discussion part also included current limitation and future perspectives. Scientifically, this study is perfect, suitable for publication. However, a revision is required before the publication.

Wherever applicable, authors need to cite recent literature reports and increase the number of references.
Some typos and need to be corrected: cell [99–101] and [91–93].
If the author used any AI tools for creating figures, that need to be mentioned in the acknowledgment section.
A specific paragraph to be provided related to the inhibitors or small organic molecules interfering with nuclear mechanics and nuclear mechanotransduction events, with a table.

Author Response

Dear Reviewer 1

Thank you very much for the helpful comments, all of which I have taken into account when editing the manuscript. They are marked in yellow in the uploaded version (see below).

Reviewer 1

Wherever applicable, authors need to cite recent literature reports and increase the number of references.

Answer: I have added new references, but since Revier 3 has already pointed out that the manuscript is too long, I have limited myself to the essentials.

Some typos and need to be corrected: cell [99–101] and [91–93].

Answer: Thank you for pointing this out. I apologize for the typos and have corrected them throughout the manuscript.

If the author used any AI tools for creating figures, that need to be mentioned in the acknowledgment section.

Answer: I did not use any AI tools to create the illustrations. I designed them all myself.

A specific paragraph to be provided related to the inhibitors or small organic molecules interfering with nuclear mechanics and nuclear mechanotransduction events, with a new table.

Answer: This is a very good suggestion, which I am pleased to follow. I have supplemented the text and inserted a new table.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Please see the attached file

Comments for author File: Comments.pdf

Author Response

Dear Reviewer 2

Thank you very much for the helpful comments, all of which I have taken into account when editing the manuscript. They are marked in yellow in the uploaded version (see below).

Reviewer 2

The review is comprehensive and conceptually ambitious. However, several structural, formatting, and mechanistic issues should be addressed to improve clarity and scientific precision.

Overloaded figure legends (Figures 2, 4, 5, 8) - all four legends should be reduced substantially and avoid conceptual repetition. Legends should describe the figure, not reexplain signaling pathways.

The legends of Figures 2, 4, 5, and 8 are excessively long and partially duplicate entire sections of the manuscript.

* **Figure 2 (pp. 8–9)**: The legend includes extended explanations of YAP/TAZ, RAR/ERα signaling, cytoskeletal organization, and ciliary mechanotransduction. Most of this is already covered in Sections 2.3–2.4.

* **Figure 4 (pp. 22–24)**: Full structural description of the LINC complex (SUN/KASH, nesprin isoforms, lamin interactions) repeats Section 5.1.

* **Figure 5 (pp. 26–27)**: Detailed Ran-GTP/NTR/NLS/NES explanation exceeds what is required for a schematic.

* **Figure 8 (pp. 37–38)**: The legend restates chromatin modification mechanisms (HDAC3, H3K9me3, H3K27me3, CpG methylation) already discussed in Section 7.2.

Answer: I have reworded the figure legends accordingly.

Formatting issues

(a) Bold references -some references appear in bold without explanation. This should be corrected for consistency.

(b) Gene nomenclature - gene symbols are not consistently italicized (e.g., *LMNA*, *Suv39H1*, *Setdb1*, *RBFOX1*, *PXN*, *VCL*, *HMGB2*). Genes should be italicized; protein names should not.

(c) Repeated abbreviation definitions - LINC, NPC, cPLA2, YAP/TAZ, H3K9me3/H3K27me3 are defined multiple times (Sections 2.5, 4.1, 5.1, 7.2, 8.1–8.2). Define once and avoid repetition.

Answer: (A) They are no longer in bold print, but in normal font. My apologies, but this was an issue with my reference program.

(B) I have corrected the gene nomenclature.

(C) LINC has been defined in the abstract and in the text, which is standard. NPC has been defined in figure legend 3 and in the manuscript text, which is also correct. cPLA2 is defined in the text and in the heading (just for simplicity), which is standard. YAP/TAZ it is now defined in the text. H3K9me2 is now defined and in the abbreviation list. H3K9me3 is defined once. I have omitted the H3K9me3/H3K27me3 repetitions in the text.

Redundancy of mechanotransduction definitions - the distinction between mechanosensing and mechanotransduction is clearly defined in Section 1 (pp. 2–4), but conceptually repeated in Sections 2.5, 4.1, and 8.1. After the initial definition, refer back instead of redefining.

Answer: I have reduced the redundancy by rewording the text accordingly.

Insufficient quantitative integration - the manuscript discusses nuclear stiffness and viscoelasticity extensively but rarely provides numerical ranges or mechanical context.

For example: Section 4.2: stiffness varies “by orders of magnitude” - no values provided; Section 7.4: protrusive forces (0.2–0.5 nN) are mentioned without comparison to nuclear yield thresholds; Section 8.2: 5 pN chromatin decondensation is cited without contextualization. Given the central role of mechanics in invasion, quantitative framing is necessary.

Answer: I agree and provide now numbers in the manuscript. I provide now stiffness values in section 4.2. I also included a new part 4.4 that connects nuclear stiffness with migration and invasion.

Cancer focus needs sharpening - large parts of Sections 6–7 read as general nuclear biology without consistent linkage to invasion (ATRX/DAXX/H3.3 deposition (Section 6.2) lacks explicit connection to migration mechanics; LLPS discussion (Section 7.3) is not consistently tied to metastatic plasticity.)

The discussion of mechanomarkers could be strengthened by citing work that connects mechanical invasiveness with metastatic risk.

Answer: I have included a new subsection 4.4 that connects mechanomarker with invasion and metastasis.

The manuscript includes 11 self-citations. While potentially justified, statements primarily supported by the author’s prior work should be balanced with independent literature.

Answer: I have over 400 references, although I was only involved in 11 manuscripts, which corresponds to about 0.25%, which is very little compared to other manuscripts. During the revision, I added a few more references.

Section 8.1 states that viewing the nucleus as a single mechanosensor is simplistic, yet later concludes that the entire nucleus acts as a mechanosensor.

Answer: I agree. The traditional view of the cell nucleus is that of a mechanosensory whole, but this can be questioned. I have adjusted the headline accordingly.

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

In the present 71- page long manuscript, Claudia Tanja Mierke reviews in much detail the structure and function of cell nuclei with emphasis on mechanics. The list of references is extensive, mostly relevant. Illustrations are clarifying the text.

The following comments are meant to improve the manuscript.

One. Shortening of the text might be realized by omitting general textbook knowledge and redundant information. An example of the former can be found on L 1588 to 1595, starting with “Chromatin is the genetic material that is composed of DNA”. The latter is exemplified by a comparison between sections L619-621 and L733 – 735, respectively. Another example can be found on L L575- L593.

Two. The title anounces “cancer cell migration and invasion”, but data are limited, not critically analyzed, and references are not always relevant. This is the weak part of an otherwise solid paper.

Examples:

L 570 : Ref 122 dealing with NIH 3T3 fibroblasts and A549 human lung carcinoma-derived cells bears no relevance for migration or invasion.

L693- 700: Ref 127 is not about cancer.

L859: states “In contrast, elevated nuclear stiffness may point to enhanced fluidity of cancer cells and increased metastatic capacity based on references 205 and 206”. Ref 205, (Wolff et al., 2007) covers types of migration of cancer cells in vitro, not metastasis. Ref 206, (Niethammer, P, 2021) is a review covering “ the basic chemical and mechanical properties of nuclear components, and how these properties are thought to be utilized for mechanosensing”, as cited in the abstract. Cancer cells are mentioned together with other cell types, but neither invasion nor metastasis is discussed.

L 1338: “Thus, the MAT transition, which is frequently observed in melanoma and other cancers, enhances the plasticity of cells and enables more efficacious movement and metastasis even under challenging circumstances like confinements”. On line 1329 we find a putative support for this satetement in the form of reference 140. This paper by the author of the present manuscript (Mierke C T , 2019) is a review of physical concepts and hypotheses pleading in favour of “including biomechanical properties of cells, cell clusters and tissues and their microenvironment to understand mechano-regulatory processes within cancer cells and the entire organism”, useful but not presenting evidence for the statement on line 1338. We note a good deal of redundancy between the latter and the present manuscript.

L1483: Ref 304 (Hearst S et al., 2009) concerns HeLa cells and WI-38 cells, termed transformed and primary cell lines with doubtful relevance for cancer and none for invasion. Similarly, Ref 305 (Spector D. et al., 1992) uses “transformed cells, immortal cells, and cells of defined passage number” of different nature and origine and concludes that “the organization of snRNPs within the mammalian cell nucleus is a reflection of the physiology of the cell that may change upon transformation or immortalization” with no relevance for invasion.

L1543: General statements such as “Abnormalities in the structure and functioning of nuclear speckles are closely linked to certain diseases, like cancer.” are too vague.

L671 – 678: This “summary” is not underpinned by the abovementioned data .

Author Response

Dear Reviewer 3

Thank you very much for the helpful comments, all of which I have taken into account when editing the manuscript. They are marked in yellow in the uploaded version (see below).

Reviewer 3

The following comments are meant to improve the manuscript.

Answer: I have made some adjustments to the text. It is difficult for me to satisfy the conflicting demands of all three reviewers. The other two reviewers even requested additions, which I have kept to a minimum. I have slightly reworded these sections, but this information is still necessary for readers, who mostly come from different disciplines.

Two. The title announces “cancer cell migration and invasion”, but data are limited, not critically analyzed, and references are not always relevant. This is the weak part of an otherwise solid paper.

Answer: It is difficult to cover the entire subject area comprehensively in a single manuscript, so I have included many references that offer interested readers the opportunity to obtain further information on less important aspects. I have explained the link between cancer cell migration and invasion using nuclear technology in more detail in one section.

Examples:

L 570: Ref 122 dealing with NIH 3T3 fibroblasts and A549 human lung carcinoma-derived cells bears no relevance for migration or invasion.

Answer: Thank you for pointing it out. I have amended the manuscript text and added additional information. I have also included several additional references.

L693- 700: Ref 127 is not about cancer.

Answer: This article should be cited here in general terms. I have adapted the text accordingly. I have included a new subsection 4.4 that connects the nuclear mechanics to cancer cell migration and invasion.

L859 (is 895): states “In contrast, elevated nuclear stiffness may point to enhanced fluidity of cancer cells and increased metastatic capacity based on references 205 and 206”. Ref 205, (Wolff et al., 2007) covers types of migration of cancer cells in vitro, not metastasis. Ref 206, (Niethammer, P, 2021) is a review covering “the basic chemical and mechanical properties of nuclear components, and how these properties are thought to be utilized for mechanosensing”, as cited in the abstract. Cancer cells are mentioned together with other cell types, but neither invasion nor metastasis is discussed.

Answer: I agree and have reworded it and inserted the citations elsewhere. I have also added new references. In addition, I have added a new section 4.4, which now addresses this issue.

L 1338: “Thus, the MAT transition, which is frequently observed in melanoma and other cancers, enhances the plasticity of cells and enables more efficacious movement and metastasis even under challenging circumstances like confinements”. On line 1329 we find a putative support for this statement in the form of reference 140. This paper by the author of the present manuscript (Mierke C T, 2019) is a review of physical concepts and hypotheses pleading in favour of “including biomechanical properties of cells, cell clusters and tissues and their microenvironment to understand mechano-regulatory processes within cancer cells and the entire organism”, useful but not presenting evidence for the statement on line 1338. We note a good deal of redundancy between the latter and the present manuscript.

Answer: I cited a new reference for this statement “Thus, the MAT transition, which is frequently observed in melanoma and other cancers, enhances the plasticity of cells and enables more efficacious movement and metastasis even under challenging circumstances like confinements”. The Mierke C T, 2019 is cited as a review article. I also cited now two new references for the piston-migration mode.

Answer: I have included this important point and reworded the text and also included another reference. I reworded the point with the invasion as a hypothesis.

L1543: General statements such as “Abnormalities in the structure and functioning of nuclear speckles are closely linked to certain diseases, like cancer.” are too vague.

Answer: I reworded it and provide an example. I also included a new citation.

L671 – 678: This “summary” is not underpinned by the abovementioned data.

Answer: I fully agree and reworded it. It is now formulated as an outlook and connection to the next section.

Author Response File: Author Response.pdf