The chloride cotransporter NKCC1 regulates self-renewal of hippocampal neural stem cells via the transcription factor Sox11

Blank, A.; Adhikarla, S.; Broesske, J.; Kaiser, L.; Rippe, M.; Haase, M.; Groth, M.; Stein, G.; Andrae, T.; Mesterheide, J.; Fleischer, A.-L.; Flossmann, T.; Schaeffner, I.; Sock, E.; Kirmse, K.; Huebner, C. A.; Holthoff, K.; Schmeer, C.; Lie, D. C.; Keiner, S.

doi:10.1101/2025.07.01.662334

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bioRxiv · 10.1101/2025.07.01.662334

The chloride cotransporter NKCC1 regulates self-renewal of hippocampal neural stem cells via the transcription factor Sox11

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Abstract

The GABAergic-mediated depolarization plays a key role in controlling stem cell fate and neurogenesis within the dentate gyrus of the hippocampal formation. This depolarization effect is highly dependent on the balance between the chloride co-transporters NKCC1 and KCC2. It is not known how changes in NKCC1 modulate the fate of Nestin-positive stem cells (NSCs) in the hippocampus during adult neurogenesis. In our study, we demonstrate that a knockout of Nkcc1 in NSCs increase their proliferation by symmetric self-renewal and expand the stem cell pool. Using single-cell RNA sequencing, we identified Sox11 as a key transcription factor that is significantly downregulated following Nkcc1 knockout. In agreement with this finding, we found that Sox11 knockout enhances proliferation and self-renewal of NSCs, which is also marked by an increase in symmetric stem cell division. Based on these findings, we propose that altering Nkcc1 expression in NSCs shifts their fate from neurogenesis towards self-renewal via Sox11 regulation. Furthermore, we observed that NKCC1 levels decline in NSCs during aging, which correlates with a further increase in self-renewal. Our data strongly suggest that the age-related decline in NKCC1 levels promote symmetric division and self-renewal contributing to the age-dependent decrease in neurogenesis. NKCC1 via Sox11 is a key regulator of NSCs fate decision, critically balancing self-renewal and neuronal differentiation in the adult hippocampus. HighlightsNKCC1 is a key factor in regulating the Nestin-positive neural stem cell symmetric self-renewal. Sox11 is a downstream effector of NKCC1 in Nestin-positive stem cells and is involved in expansion of the stem cell pool. Aging reduces NKCC1 expression in Nestin-positive stem cells and increases their self-renewal. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=170 HEIGHT=200 SRC="FIGDIR/small/662334v1_ufig1.gif" ALT="Figure 1000"> View larger version (38K): org.highwire.dtl.DTLVardef@1babdcforg.highwire.dtl.DTLVardef@1038bfborg.highwire.dtl.DTLVardef@1db105corg.highwire.dtl.DTLVardef@10f2cde_HPS_FORMAT_FIGEXP M_FIG C_FIG

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Blank, A., Adhikarla, S., Broesske, J., Kaiser, L., Rippe, M., Haase, M., Groth, M., Stein, G., Andrae, T., Mesterheide, J., Fleischer, A.-L., Flossmann, T., Schaeffner, I., Sock, E., Kirmse, K., Huebner, C. A., Holthoff, K., Schmeer, C., Lie, D. C., Keiner, S.. 2025-07-04. The chloride cotransporter NKCC1 regulates self-renewal of hippocampal neural stem cells via the transcription factor Sox11. https://doi.org/10.1101/2025.07.01.662334

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