I am very excited to share this preprint which has been my main postdoc project! We identified leupaxin as a negative regulator of durotaxis via paxillin-FAK inhibition in MDA-MB-231 brain and bone tropic metastatic variants. www.biorxiv.org/content/10.6...
Congratulations Giulia!
Thanks Helen!! :)
How do cells adapt their volumes when they proliferate and form a multicellular mass in 3D? If you have wondered about this or work with 3D in-vitro systems, then you will be happy to read our paper now on bioRxiv!
www.biorxiv.org/content/10.1...
See the thread 🧵 below for more information.
Thanks Judith!
This forms a major part of my PhD and I would be happy to get feedback on this!
Additionally, we see changes in mechanical properties (through Brillouin microscopy) of cells along with these volume changes. Single cells (both in the case of proliferation and invasion) were more compressible compared to cells in multicellular structures. Such a coupling is seen in many cases.
We rather propose that the multicellular context of the cell determines how the cell modulates its volume. We also have some patient data (H&E sections) to push that point a bit further.
Let us know what you think about this!
Han et al. had proposed that these volume changes during invasion are due to a compressive stress gradient (from centre to periphery). This drives intracellular flow of water between cells, through gap junctions. We do not see any gradient of volume in our spheroids (like most people in the field).
Here we had a single cell becoming a multicellular entity. In invasion, it is the opposite - a multicellular entity gives rise to 'single' cells. We (along with Han et al. 2020 Nat Phy) see the same trend in this case. Invading cells are bigger compared to cells at the centre of the spheroid.
Thus, we propose that changes in water content (leading to mass density changes) along with changes in cell cycle distribution, together, lead to the drastic difference in the nuclear volumes between single cells and multicellular structures.
This was also suggested by looking at the refractive index of cells. RI scales with the mass density of cells. We observed higher RI for multicellular structures compared to single cells. So when cells were released from this context, they osmotically balanced themselves and increased in volume.
We then released single cells at different timepoints (by breaking down the gel and the clusters/spheroids) and looked at their nuclear volumes. We observed that when cells were released from a multicellular context, they (probably) took in water and increased their nuclear volume.
By blocking cells in S/G2 phase with a CDK1 inhibitor, we were able to rescue the nuclear volumes of small clusters. But this increased nuclear volume still did not reach the single cell level. This hinted that another mechanism was at play.
As single cells formed these multicellular structures, there was an accumulation of smaller G1 cells over time. Light sheet microscopy allowed us to show that this was due to prolongation of the G1 phase of the cell cycle, while S/G2 phase length did not change much over time.
Then why are the nuclear volumes decreasing? We did some coarse timelapse microscopy to tackle that. We saw that cells were growing over the cell cycle and could become as big as their mother cell. So, there was no inhibition of growth in this system. But this hinted at us to look at the cell cycle.
You might guess that nuclear volumes might decrease due to compressive stress on the encapsulated spheroids. We tested that using the gel system. By changing the degradability and stiffness of the gel, we can alter the compressive stress on the spheroids. But that did not affect the nuclear volumes.
Our model system - single cells in 3D hydrogels that proliferated to form clonal spheroids. We were surprised to see drastic reductions in nuclear volumes upon nuclear segmentation in 3D. Below the data is for MCF-7 but we see this for multiple cell lines - PANC1, MCF10A, HeLa.
Let's start with some motivation! 💪
Cells tightly regulate their volumes, especially in the face of external perturbations like osmolarity, confinement, etc. Very few studies have looked at such relationships in 3D, especially in the context of multicellularity - during proliferation and invasion.
How do cells adapt their volumes when they proliferate and form a multicellular mass in 3D? If you have wondered about this or work with 3D in-vitro systems, then you will be happy to read our paper now on bioRxiv!
www.biorxiv.org/content/10.1...
See the thread 🧵 below for more information.
Malcolm Gladwell’s Revisionist History podcast is the best thing ever!
All the best! 💪🏻
Congratulations!
Thanks for organising this amazing conference! 😬
It’s the #filopodia, stupid! 🚀
We just released our new study uncovering how filopodia and hydrostatic #pressure contribute to the regulation of #membrane #tension.
👉 Check it out on bioRxiv: biorxiv.org/content/10.1...
#cellbiology #biophysics #actin #mechanobiology
Want to get started with analysing your cells/spheroids/organoids in 3D matrices? Check out our review about imaging, biomechanical and biochemical phenotyping in 3D @cp-trendscellbio.bsky.social
www.sciencedirect.com/science/arti...
Want to get started with analysing your cells/spheroids/organoids in 3D matrices? Check out our review about imaging, biomechanical and biochemical phenotyping in 3D @cp-trendscellbio.bsky.social
www.sciencedirect.com/science/arti...
I wrote this some time back - ‘PhD is for training’
🚨THREE fully-funded PhD positions available! 2 in our lab @mcb-sheffield.bsky.social and 1 with our collaborators in Leeds.
Three exciting but very different projects in cancer cell biology, biophysics & chemical/structural biology.
👇 full details in 🧵
