Investigating Tunnelling Nanotube Mediated Intercellular Transfer of Mitochondria in Response to Mitochondrial Import Failure

Abstract

Protein import into mitochondria is essential for normal cellular function. Impaired
import results in defective mitochondrial respiration and depletion of the major cellular
ATP source, and this is particularly damaging to cells with high energetic demands
such as cancer cells and neurons. Cells activate stress response pathways to
overcome acute import impairment, but prolonged impairment leads to disease.
Previously, our lab has shown that chronic import failure is rescued by intercellular
transfer of mitochondria via tunnelling nanotubes (TNTs), actin-based cellular
protrusions that mediate membrane continuity between connected cells. However,
how this intercellular transfer mediates cellular rescue is currently unknown.
Moreover, more generally, the mechanisms underpinning TNT-mediated transfer, the
fate of the transferred mitochondria, and the relevance of these mechanisms in vivo,
are still poorly understood.
Here, I used live imaging, correlative light and electron microscopy (CLEM) and flow
cytometry approaches to characterise transport of mitochondria through TNTs and
interrogate the product of intercellular mitochondrial transfer: cells containing
exogenous mitochondria. I show that TNTs facilitate both the uptake of healthy
mitochondria into import-compromised cells and expulsion of mitochondria with
blocked import sites into healthy cells. I found that exported import-defective
mitochondria have a small and spherical morphology and undergo conventional
degradation while, unexpectedly, imported healthy mitochondria are sequestered into
a large ‘mitochondrial body’ in import-defective cells for eventual degradation.
Additionally, I used flow cytometry to show that intercellular mitochondrial transfer
occurs under basal conditions and is upregulated in response to mitochondrial stress.
My data suggests intercellular transfer of mitochondria and mitophagy collaborate to
eliminate defective mitochondria.
Finally, I made zebrafish (Danio rerio) lines to investigate the impact of chronic
mitochondrial protein import failure in vivo. This preliminary data highlights the
potential of zebrafish as a in vivo model for investigating the impact of mitochondrial
import inhibition and observation of intercellular mitochondrial transfer in vivo.
Overall, the work presented in this thesis provides an insight into how TNT-mediated
intercellular mitochondrial transfer mediates rescue of chronic mitochondrial import
inhibition. Furthermore, my data suggests the involvement of this phenomenon in
mitochondrial quality control.

Date of Award	20 Jan 2026
Original language	English
Awarding Institution	University of Bristol
Supervisor	Ian R Collinson (Supervisor)