An oncogenic isoform of septin 9 promotes the formation of juxtanuclear invadopodia by reducing nuclear deformability

Background.

One of the big challenges for cancer cells is to escape the original tumor to invade a new tissue. The formation of invadopodia allows cancer cells to leave the original tumor [1]. This process involves mostly F-actin core – composed of TKS5 [2] and MT1-MMP [3] which is responsible  for matrix degradation. Interestingly, the invadopodia begin mostly under the nucleus from where they can deformed it, suggesting that the actin core is physically linked to the nucleus [4].

Another challenge for the cancer cells is to go through the extracellular matrix (ECM) with a massive and rigid organelle like the nucleus. Cells must “find” ways to passing the ECM without damaging the nucleus too much. There are several possibilities. One is to make the nuclear envelope softer and another one is matrix degradation. However, these mechanisms remain little understood during invadopodia formation.

Septin, a GTP-binding protein interacting with the cytoskeleton [5, 6], could also be involved in invadopodia formation because it has been shown that Spet9-i1 overexpression enhances cancer cells invasion and metastasis [7].

The question now is:

Is there a link between the nucleus and septins during invadopodia formation?

The preprint “An oncogenic isoform of septin 9 promotes the formation of juxtanuclear invadopodia by reducing nuclear deformability” proposes a mechanism in which Sept9_i1 protects the nucleus against deformability during the early steps of cell invasion. To conduct this study, the authors used breast cancer cells containing a high copy number of sept9_i1 in 2D and 3D ECM assay.  The 3D assay involved plating cells on permeable Matrigel and a laminin-coated Transwell membrane. They used EGF to stimulate invadopodia formation.

 

Key findings.

• Sept9_i1 dynamics in invadopodia.

Sept9_i1 is specifically required for invadopodia formation because its inhibition by short hairpin RNA (shRNA) prevents gelatin degradation and invadopodia.

The authors have observed a dynamic localization of Sept9_i1 during invadopodia formation. At the beginning of invadopodia stimulation (12 hrs) spet9_i1 is located in proximal part (12 of the invadopodia after which it can be found either throughout or only in the distal part (24 hrs).

Since TKS5 and cortactin are both key factor to invadopodia formation, the authors looked whether sept9_i1 could impact these two factors. They showed that sept9_i1 plays a role in TKS5 and cortactin cluster formation which are located at the cell membrane and around the nucleus. Indeed Sept9_i1 knockdown affects TKS5 and cortactin subcellular organization.

Interestingly, in this first part the authors show that sept9-i1 is important for invadopodia formation and maturation through its association with TKS5.

 

• Sept9_i1: a bodyguard for the nucleus.

Knowing that invadopodia formation starts under the nucleus, spet9-i1 could be located near to it. By using super-resolution confocal microscopy, the authors found that Sept9_i1 is in the ventral part and in basolateral part of the nucleus. Indeed, Sept9-i1 colocalized with LaminA/C and seems to be intercalated between nucleoporins. However, why is sept9_i1 appended to the nuclear envelope? Surprisingly, Sept9_i1 knockdown led to a nucleus with many wrinkles enriched in laminA/C and nucleoporins compared to control and sept9_i2. The authors speculated that Sept9_i1 could act like lamins and control nuclear rigidity outside it. To confirm this hypothesis, they overexpressed lamin A in Sept9_i1 depleted cells. Lamin A rescued the nuclear shape and the TSK5 cluster numbers. Therefore, the formation of invadopodia seems to be linked to nuclear mechanical properties. Indeed, the authors propose a new mechanism where sept9_i1 mechanically controls the rigidity of the nucleus and invadopodia formation.

In this study, the authors highlighte a new invadopodia factor and a new nuclear mechanobiology factors: Sept9_i1. They conclude that Sept9_i1 allows a functional connection between the nucleus and invadopodia formation. It acts as a nuclear morphology protector, which is important for invadopodia formation itself. Indeed, they propose that a rigid nucleus is essential to allow actin polymerization and support forces generated by the formation of the strong actin core at the plasma membrane under the nucleus.  Finally, invadopodia formation at the juxtanuclear plasma membrane allows ECM degradation to promote nucleus translocation without drastic deformation. Sept9_i1 seems to be a new metastatic factor in breast cancer which is linked to nuclear mechanics.

 

Why I picked this preprint.

Understanding the nuclear mechanics during cancer progression and how cancer cells can manage invasion with this “big ball” inside remain essential questions. This preprint offers a new answer that may advance our knowledge of the cell invasion mechanisms and nuclear mechanics during metastasis.

I am really interested in the non-genomic role of the nucleus, nuclear physical properties and nuclear-mechanotransduction. In this preprint, I really like the idea that there can be a protective outer nuclear barrier (Sept9_i1) to complement the inner nuclear barrier (lamins). Both of them control the nuclear rigidity to avoid nuclear deformation during cancer metastasis and importantly gene expression modification.

 

Questions for the authors.

Q1. Have you looked at the Sept9_i1 dynamics during nuclear translocation?

Q2. Is it possible that the “external wall” (Sept9_i1) and the “internal wall” interact to enhance the nuclear rigidity? As you mentioned in your discussion Sept9_i1 could interact with the LINC complex. Consequently, it will be very interesting for future studies to see if LINC complex depletion affects spetin organization and nuclear shape. Or if overexpression of it can also rescue the phenotype. And to observe if sept9_i1 colocalizes with the LINC complex. However, in some types of breast cancer the LINC complex is downregulated [8]. As a result, I am wondering if the LINC complex could be involved in this process at least in some types of breast cancer. We can also speculate that if the LINC complex is downregulated, the oncegene sept9_i1 could compensate for its loss. I wonder what is your thoughts are these fundamental questions.

Q3. Did you try to knock down lamins to observe the sept9_i1 dynamics?

Q4. Your study is about cancer cell behavior during invasion. Do you think this kind of septin organization can be observed in normal cells, and play a similar role in nuclear rigidity? For example, for cells which have to support lot of tension. And in your opinion, could this mechanisms also exist during physiological cell migration? For example, during development.

 

References.

1. Linder, S., et al., Mechanisms and roles of podosomes and invadopodia. Nat Rev Mol Cell Biol, 2023. 24(2): p. 86-106.
2. Blouw, B., et al., A role for the podosome/invadopodia scaffold protein Tks5 in tumor growth in vivo. Eur J Cell Biol, 2008. 87(8-9): p. 555-67.
3. Ferrari, R., et al., MT1-MMP directs force-producing proteolytic contacts that drive tumor cell invasion. Nat Commun, 2019. 10(1): p. 4886.
4. Revach, O.Y., et al., Mechanical interplay between invadopodia and the nucleus in cultured cancer cells. Sci Rep, 2015. 5: p. 9466.
5. Spiliotis, E.T. and K. Nakos, Cellular functions of actin- and microtubule-associated septins. Curr Biol, 2021. 31(10): p. R651-R666.
6. Benoit, B., C. Poüs, and A. Baillet, Septins as membrane influencers: direct play or in association with other cytoskeleton partners. Front Cell Dev Biol, 2023. 11: p. 1112319.
7. Connolly, D., et al., Septin 9 isoform expression, localization and epigenetic changes during human and mouse breast cancer progression. Breast Cancer Res, 2011. 13(4): p. R76.
8. Matsumoto, A., et al., Global loss of a nuclear lamina component, lamin A/C, and LINC complex components SUN1, SUN2, and nesprin-2 in breast cancer. Cancer Med, 2015. 4(10): p. 1547-57.

 

Tags: invadopodia, nucleus, septin

doi: https://doi.org/10.1242/prelights.35224

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