Ral-GTPase interacts with the beta1 subunit of Na+/K+-ATPase and is activated upon inhibition of the Na+/K+ pump.

Abstract

Na+/K+-ATPase functions as both an ion pump and a signal transducer. Cardiac glycosides partially inhibit Na+/K+-ATPase, causing activation of multiple interrelated growth pathways via the Na+/K+-ATPase/c-Src/epidermal growth factor receptor complex. Such pathways include Ras/MEK/ERK and Ral/RalGDS cascades, which can lead to cardiac hypertrophy. In search of novel Ral-GTPase binding proteins, we used RalB as the bait to screen a human testes cDNA expression library using the yeast 2-hybrid system. The results demonstrated that 1 of the RalB interacting clones represented the C-terminal region of the beta1 subunit of Na+/K+-ATPase. Further analysis using the yeast 2-hybrid system and full-length beta1 subunit of Na+/K+-ATPase confirmed the interaction with RalA and RalB. In vitro binding and pull-down assays demonstrated that the beta1 subunit of Na+/K+-ATPase interacts directly with RalA and RalB. Ral-GTP pull-down assays demonstrated that short-term ouabain treatment of A7r5 cells, a rat aorta smooth muscle cell line, caused activation of Ral GTPase. Maximal activation was observed 10 min after ouabain treatment. Ouabain-mediated Ral activation was inhibited upon the stimulation of Na+/K+-ATPase activity by Ang II. We propose that Ral GTPase is involved in the signal transducing function of Na+/K+-ATPase and provides a possible molecular mechanism connecting Ral to cardiac hypertrophy during diseased conditions.

• Authors: Rajinder P Bhullar, Richard R Clough, Juddy Kanungo, Sherif M Elsaraj, Ognjen Grujic
• Journal: Canadian journal of physiology and pharmacology (Can J Physiol Pharmacol) 2007 Mar-Apr Vol. 85 Issue 3-4 Pg. 444-54 ISSN: 0008-4212 [Print] Canada
• PMID: 17612654 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Atp1b1 protein, rat
• DNA, Complementary
• Ralb protein, rat
• Recombinant Proteins
• Angiotensin II
• Ouabain
• Glutathione Transferase
• Rala protein, rat
• ral GTP-Binding Proteins
• Sodium-Potassium-Exchanging ATPase

Topics

• Angiotensin II (pharmacology)
• Animals
• Cell Line
• DNA, Complementary (genetics)
• Glutathione Transferase (metabolism)
• Humans
• Male
• Ouabain (pharmacology)
• Plasmids
• Rats
• Recombinant Proteins (metabolism)
• Sodium-Potassium-Exchanging ATPase (antagonists & inhibitors, genetics, metabolism)
• Testis (metabolism)
• Two-Hybrid System Techniques
• Yeasts (genetics)
• ral GTP-Binding Proteins (genetics, metabolism)