Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 112, Issue 24, Pages E3104-E3113Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1421290112
Keywords
electrolyte; cell volume; hypertonicity; membrane trafficking; mitochondria
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Funding
- Swiss National Science Foundation [31003A_138408/1]
- Fondation Schmidheiny
- Sir Jules Thorn Charitable Overseas Trust
- Swiss National Science Foundation (SNF) [31003A_138408] Funding Source: Swiss National Science Foundation (SNF)
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Cell volume homeostasis is vital for the maintenance of optimal protein density and cellular function. Numerous mammalian cell types are routinely exposed to acute hypertonic challenge and shrink. Molecular crowding modifies biochemical reaction rates and decreases macromolecule diffusion. Cell volume is restored rapidly by ion influx but at the expense of elevated intracellular sodium and chloride levels that persist long after challenge. Although recent studies have highlighted the role of molecular crowding on the effects of hypertonicity, the effects of ionic imbalance on cellular trafficking dynamics in living cells are largely unexplored. By tracking distinct fluorescently labeled endosome/vesicle populations by live-cell imaging, we show that vesicle motility is reduced dramatically in a variety of cell types at the onset of hypertonic challenge. Live-cell imaging of actin and tubulin revealed similar arrested microfilament motility upon challenge. Vesicle motility recovered long after cell volume, a process that required functional regulatory volume increase and was accelerated by a return of extracellular osmolality to isosmotic levels. This delay suggests that, although volume-induced molecular crowding contributes to trafficking defects, it alone cannot explain the observed effects. Using fluorescent indicators and FRET-based probes, we found that intracellular ATP abundance and mitochondrial potential were reduced by hypertonicity and recovered after longer periods of time. Similar to the effects of osmotic challenge, isovolumetric elevation of intracellular chloride concentration by ionophores transiently decreased ATP production by mitochondria and abated microfilament and vesicle motility. These data illustrate how perturbed ionic balance, in addition to molecular crowding, affects membrane trafficking.
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