Ouabain‐regulated phosphoproteome reveals molecular mechanisms for Na + , K + ‐ATPase control of cell adhesion, proliferation, and survival

Authors:
Open Access
Publisher website
Google Scholar
Abstract
The ion pump Na + , K + –ATPase (NKA) is a receptor for the cardiotonic steroid ouabain. Subsaturating concentration of ouabain triggers intracellular calcium oscillations, stimulates cell proliferation and adhesion, and protects from apoptosis. However, it is controversial whether ouabain‐bound NKA is considered a signal transducer. To address this question, we performed a global analysis of protein phosphorylation in COS‐7 cells, identifying 2580 regulated phosphorylation events on 1242 proteins upon 10‐ and 20‐min treatment with ouabain. Regulated phosphorylated proteins include the inositol triphosphate receptor and stromal interaction molecule, which are essential for initiating calcium oscillations. Hierarchical clustering revealed that ouabain triggers a structured phosphorylation response that occurs in a well‐defined, time‐dependent manner and affects specific cellular processes, including cell proliferation and cell‐cell junctions. We additionally identify regulation of the phosphorylation of several calcium and calmodulin–dependent protein kinases (CAMKs), including 2 sites of CAMK type II‐γ (CAMK2G), a protein known to regulate apoptosis. To verify the significance of this result, CAMK2G was knocked down in primary kidney cells. CAMK2G knockdown impaired ouabain‐dependent protection from apoptosis upon treatment with high glucose or serum deprivation. In conclusion, we establish NKA as the coordinator of a broad, tightly regulated phosphorylation response in cells and define CAMK2G as a downstream effector of NKA.—Panizza, E., Zhang, L., Fontana, J. M., Hamada, K., Svensson, D., Akkuratov, E. E., Scott, L., Mikoshiba, K., Brismar, H., Lehtiö, J., Aperia, A. Ouabain‐regulated phosphoproteome reveals molecular mechanisms for Na + , K + –ATPase control of cell adhesion, proliferation, and survival. FASEB J. 33, 10193–10206 (2019). www.fasebj.org
Funding Information
  • Novo Nordisk

This publication has 72 references indexed in Scilit: