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Title: The flip side of the arabidopsis type I proton-pumping pyrophosphatase (AVP1): Using a trans-membrane H+ gradient to synthesize pyroposphate

Author
item SCHOLZ-STARKE, JOACHIM - Consiglio Nazionale Delle Ricerche
item PRIMO-PLANTA, CECILIA - Children'S Nutrition Research Center (CNRC)
item YANG, JIAN - Children'S Nutrition Research Center (CNRC)
item KANDEL, RAJU - Arizona State University
item GAXIOLA, ROBERTO - Arizona State University
item HIRSCHI, KENDAL - Children'S Nutrition Research Center (CNRC)

Submitted to: Journal of Biological Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/3/2018
Publication Date: 1/25/2019
Citation: Scholz-Starke, J., Primo-Planta, C., Yang, J., Kandel, R., Gaxiola, R.A., Hirschi, K.D. 2019. The flip side of the arabidopsis type I proton-pumping pyrophosphatase (AVP1): Using a trans-membrane H+ gradient to synthesize pyroposphate. Journal of Biological Chemistry. 294(4):1290-1299.

Interpretive Summary: Nutrient regulation is vital for normal plant growth and development. Some proteins function in multiple tissues to regulate the nutritional content of a plant. In this work we use various experimental approaches both in a test tube and in plants to demonstrate that a particular plant protein can function both to synthesis and destroy a specific metabolite. We postulate that both functions contribute to modulating and promoting crop yield. We propose a model where the synthase activity maintains the metabolite when cells encounter a difficult environment where there is high energy demands and/or low oxygen. The degradation mode may be used when energy supplies are high and the plant is able to get energy from photosynthesis. We suggest that the ability to "flip" the function of this protein between synthesis and degradation is an important aspect of modulating crop yield.

Technical Abstract: Energy partitioning and plant growth are mediated in part by a type I H+-pumping pyrophosphatase (H+-PPase). A canonical role for this transporter has been demonstrated at the tonoplast where it serves a job-sharing role with V-ATPase in vacuolar acidification. Here, we investigated whether the plant H+-PPase from Arabidopsis also functions in "reverse mode" to synthesize PPi using the transmembrane H+ gradient. Using patch-clamp recordings on Arabidopsis vacuoles, we observed inward currents upon Pi application on the cytosolic side. These currents were strongly reduced in vacuoles from two independent H+-PPase mutant lines (vhp1-1 and fugu5-1) lacking the classical PPi-induced outward currents related to H+ pumping, whereas they were significantly larger in vacuoles with engineered heightened expression of the H+-PPase. Current amplitudes related to reverse-mode H+ transport depended on the membrane potential, cytosolic Pi concentration, and magnitude of the pH gradient across the tonoplast. Of note, experiments on vacuolar membrane–enriched vesicles isolated from yeast expressing the Arabidopsis H+-PPase (AVP1) demonstrated Pi-dependent PPi synthase activity in the presence of a pH gradient. Our work establishes that a plant H+-PPase can operate as a PPi synthase beyond its canonical role in vacuolar acidification and cytosolic PPi scavenging. We propose that the PPi synthase activity of H+-PPase contributes to a cascade of events that energize plant growth.