Overexpression of a poplar two-pore K+ channel enhances salinity tolerance in tobacco cells

Populus euphratica is a plant model intensively studied for elucidating physiological and molecular mechanisms of salt tolerance in woody species. Several studies have shown that vacuolar potassium (K+) ion channels of the two-pore K+ (TPK) family play an important role in maintaining K+ homeostasis. Here, we cloned a putative TPK channel gene from P. euphratica, termed PeTPK. Sequence analysis of PeTPK1 identified the universal K-channel-specific pore signature, TXGYGD. Over-expression of PeTPK1 in tobacco BY-2 cells improved salt tolerance, but did not enhance tolerance to hyperosmotic stress caused by mannitol (200-600 mM). After 3 weeks of NaCl stress (100 and 150 mM), PeTPK1-transgenic cells had higher fresh and dry weights than wild-type cells. Salt treatment caused significantly higher Na+ accumulation and K+ loss in wild-type cells compared to transgenic cells. During short-term salt stress (100 mM NaCl, 24-h), PeTPK1-transgenic cells showed higher cell viability and reduced membrane permeabilization compared to wild-type cells. Scanning ion-selective electrode data revealed that salt-shock elicited a significantly higher transient K+ efflux from PeTPK1-transgenic callus cells and protoplasts compared to that observed in wild-type cells and protoplasts. We concluded that salt tolerance in P. euphratica is most likely mediated through PeTPK1. We propose that, under salt stress, PeTPK1 functions as an outward-rectifying, K+ efflux channel in the vacuole that transfers K+ to the cytosol to maintain K+ homeostasis.