Anion Channel Inhibitor NPPB-Inhibited Fluoride Accumulation in Tea Plant (Camellia sinensis) Is Related to the Regulation of Ca²⁺, CaM and Depolarization of Plasma Membrane Potential.

Tian-Yuan Yang,Xian-Chen Zhang,Hong-Jian Gao,Yu-Mei Wang,Xiao-Chun Wan,Zheng-Zhu Zhang,Hong-Hong Wu

doi:10.3390/ijms17010057

Tian-Yuan Yang, Xian-Chen Zhang + Show 5 more

Open Access

https://doi.org/10.3390/ijms17010057

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Abstract

Tea plant is known to be a hyper-accumulator of fluoride (F). Over-intake of F has been shown to have adverse effects on human health, e.g., dental fluorosis. Thus, understanding the mechanisms fluoride accumulation and developing potential approaches to decrease F uptake in tea plants might be beneficial for human health. In the present study, we found that pretreatment with the anion channel inhibitor NPPB reduced F accumulation in tea plants. Simultaneously, we observed that NPPB triggered Ca2+ efflux from mature zone of tea root and significantly increased relative CaM in tea roots. Besides, pretreatment with the Ca2+ chelator (EGTA) and CaM antagonists (CPZ and TFP) suppressed NPPB-elevated cytosolic Ca2+ fluorescence intensity and CaM concentration in tea roots, respectively. Interestingly, NPPB-inhibited F accumulation was found to be significantly alleviated in tea plants pretreated with either Ca2+ chelator (EGTA) or CaM antagonists (CPZ and TFP). In addition, NPPB significantly depolarized membrane potential transiently and we argue that the net Ca2+ and H+ efflux across the plasma membrane contributed to the restoration of membrane potential. Overall, our results suggest that regulation of Ca2+-CaM and plasma membrane potential depolarization are involved in NPPB-inhibited F accumulation in tea plants.

Highlights

Fluoride (F) is phytotoxic to most plants by influencing a series of basal metabolism and enzyme activities [1]; it can be accumulated in tea plants
We reported that an anion channel inhibitor NPPB (5-nitro -2-(3-phenylpropylamino) benzoic acid) inhibited anion channels activity in tea roots and significantly reduced F accumulation in tea plants (Fuding) [7]
To investigate whether Ca2+ and CaM integrated in NPPB inhibited F accumulation in tea plants, Ca2+ flux, intracellular Ca2+ fluorescence intensity, and CaM level in tea roots were examined

Summary

Introduction

Fluoride (F) is phytotoxic to most plants by influencing a series of basal metabolism and enzyme activities [1]; it can be accumulated in tea plants. We reported that an anion channel inhibitor NPPB (5-nitro -2-(3-phenylpropylamino) benzoic acid) inhibited anion channels activity in tea roots and significantly reduced F accumulation in tea plants (Fuding) [7]. The possible mechanism involved in NPPB-inhibited F uptake in tea plants via anion channels was not yet investigated and still largely unknown. To investigate whether Ca2+ and CaM integrated in NPPB inhibited F accumulation in tea plants, Ca2+ flux, intracellular Ca2+ fluorescence intensity, and CaM level in tea roots were examined. We studied membrane potential, net H+ flux, and plasma membrane H+-ATPase activity in tea roots to investigate the possible role of regulation of membrane potential in NPPB-inhibited F accumulation in tea plants. The present study offers some potential clues to benefit the understanding of possible regulation mechanisms beyond NPPB-inhibited F accumulation in tea plants

NPPB Significantly Inhibited F Accumulation in Tea Roots and Its Whole Plant

Cultivation of Tea Plants

NPPB Treatment

Determination of F

CaM Extraction and Analysis

Membrane Potential Measurements

Data Analysis