Ion homeostasis in control of stem cell proliferation

Abstract

The ion homeostasis is critically maintained in all cells by a combined effort from various ion channels, transporters, pumps, and gap junctions. Ion fluxes are essential in excitable cells as they regulate the excitability of the cells and initiate and propagate action potentials. However, in non-excitable cells they are also involved in a number of cellular functions including proliferation, differentiation, migration, and cell volume regulation. This thesis focuses on the various mechanisms through which ion channels and transporters regulate cell cycle progression in stem cells.

Stem cells are characterized by their ability to self-renew, which entails the ability to proliferate continuously while maintaining pluripotency. Embryonic stem cells (ES cells) are pluripotent cells derived from the inner cell mass of the blastocyst and they have the ability to differentiate into any cell type originating from the three germ layers. Neural stem cells (NSCs) are multipotent adult stem cells that reside in the subventricular zone (SVZ) and differentiate to give rise to new neurons.

Previous work showed that γ-aminobutryic acid (GABA) acts on the chloride (Cl-) ion channel, GABAA receptor, to negatively regulate ES cell proliferation. We show that NSC proliferation in the adult mouse brain is regulated by a similar mechanism, where GABAA receptor activation induces the DNA damage response (DDR) pathway and the phosphorylation of histone H2AX. This results in a reduction of NSC proliferation and in long-term changes of the stem cell niche and neuronal output.

Cancer cells and stem cells share a common characteristic in their ability to proliferate extensively. We show that the main regulator of ion homeostasis in cells, the Na,K-ATPase, regulates quiescence in neuroblastoma cells. Inhibition of Na,K-ATPase with the endogenous cardiac glycoside, ouabain, induced the DDR pathway and caused a reversible cell cycle arrest. Ouabain treated cells had increased levels of the cell cycle regulator p21 and upregulated the quiescence-specific transcription factor HES1. Upon removal of ouabain, neuroblastoma cells re-entered the cell cycle and resumed proliferation.

An important role of ion homeostasis is to regulate cell volume changes over the cell cycle. The GABA signaling pathway that was shown to regulate ES cell proliferation, also altered cell volume. We found that the voltage-gated potassium (K+) channel ether-a-go-go-related gene (Erg) was differentially expressed in G1. Inhibiting Erg channel in ES cells caused a decrease in cortical stiffness and led to cell swelling followed by cell death in an apoptosis independent manner. Increasing the extracellular osmotic pressure or blocking K+ flux into ES cells resorted cell viability, suggesting that Erg channel inhibition caused cell death by increasing intracellular osmotic pressure by blocking K+ efflux from ES cells.

In summary, we show that ion homeostasis, which is maintained by ion channels and transporters, is critical for cell cycle progression in proliferating cells through various mechanisms, and that perturbation in ion homeostasis results in disrupted cell proliferation.