But don’t take just our word for it, check out the companion paper from the @septinlab.bsky.social lab: doi.org/10.64898/202...
If you want more details be sure to read the paper in full!: doi.org/10.64898/202...
From this we conclude that septins are helping to buffer mechanical forces on the nucleus, keeping it from bursting or getting damaged.
But what does this mean for the cell? Well we went back to our compression assay and pressed on these SEPT7KD cells. We used an NLS marker to keep an eye on the nucleus. Loss of septins resulted in significantly increased rupture of the nucleus under compression!
Why might septins respond this way? We decided to knockdown SEPT7 and see what happens. Septins form heteromeric subunits, and loss of SEPT7 leads to reduced expression of other isoforms too. Our SEPT2-Halo cells showed that the KD impairs formation of subnuclear fibers.
To make sure that this was independent of the nucleus we went back to our cytoplasts and added large glass beads into the cells after removing the nucleus. The beads were still sufficient to cause septin filaments to form! This is a mechanical response!
The nucleus obviously does many things, but it’s also just big. To determine whether septins were responding to biochemical or mechanical signals we tried adding in large glass beads to our fibroblasts. In response, the cells formed beautiful networks of filaments under the beads!
These experiments all suggested that the nucleus was playing a role - so we took it out! Enucleated cytoplasts lost the subnuclear network of septin (and actin) filaments.
Septin recruitment wasn’t only occurring when the cells squeezed between pillars. Instead we found that septins accumulated along the cortex whenever the nucleus was pushed up against the PDMS, regardless of the curvature of the PDMS.
Next we sought to test whether forces generated by the cell were strong enough to elicit a similar response. We used a microfluidic device designed by @lammerdinglab.bsky.social and saw septins accumulate around the nucleus as the cells squeezed between pillars.
Septins have been seen accumulating in immune cells during confined migration, so we wondered what would happen in fibroblasts. We used the dynamic confiner device developed by the Piel lab and tried compressing our cells. Septins clearly accumulate around the region of the nucleus.
Looking a little closer with the help of the @aicjanelia.bsky.social we used iPALM to see where septins were in relation to the actin. While they colocalize in the X-Y plane, septins are actually consistently below the actin stress fibers, closer to the membrane.
Watching as this fibroblast migrates, we can see that these subnuclear stress fibers move with the nucleus as the cell migrates.
Septins are another class of cytoskeletal filaments and maybe most known for their role in cell division, but they do lots of other stuff too! In interphase, septins prominently localize to a patch of subnuclear stress fibers.
So excited to share the preprint with the bulk of my PhD work in the Beach @myosincity.bsky.social and Oakes @pwoakes.bsky.social Labs at Loyola University Chicago! We investigate the structure, recruitment, and function of septin networks around the nucleus. A thread:
doi.org/10.64898/202...
@maggieutgaard.bsky.social from @pwoakes.bsky.social and @myosincity.bsky.social labs with a killer talk on the coupling of the nucleus to septins in response to mechanical stress.
#SeptSRC
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