Expression of the high-affinity K+ transporter 1 (PpHKT1) gene from almond rootstock 'Nemaguard' improved salt tolerance of transgenic Arabidopsis

Authors: KAUNDAL, AMITA - University Of California - Cooperative Extension Service; Sandhu, Devinder; DUENAS, MARCO - University Of California; Ferreira, Jorge

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/18/2019
Publication Date: 3/26/2019
Citation: Kaundal, A., Sandhu, D., Duenas, M., Ferreira, J.F. 2019. Expression of the high-affinity K+ transporter 1 (PpHKT1) gene from almond rootstock 'Nemaguard' improved salt tolerance of transgenic Arabidopsis. PLoS One. 14(3):e0214473. https://doi.org/10.1371/journal.pone.0214473.
DOI: https://doi.org/10.1371/journal.pone.0214473

Interpretive Summary: Salinity is a major abiotic stress with severe implications for the future of agriculture worldwide. Reduced availability of fresh water is forcing almond farmers to use alternate or degraded waters. As almond is sensitive crop to salinity, high concentration of salt in degraded waters is a big matter of concerns for the growers. In almonds, rootstock plays a major role in the success of a variety. Although, significant variation has been reported in different almond rootstocks, the understanding of genetic mechanism regulating salt tolerance is lacking. Plants use several different approaches to avoid or mitigate salt stress. As leaves are more sensitive to salt as compared to roots, one strategy used by plants is to stop or slow-down movement of ions from root to leaves. High affinity K+ Transporter 1 (HKT1) is known to play role regulating movement of ions to leaves. However, the HKT1 gene has not been characterized in almond rootstocks. In this investigation we have isolated and cloned the HKT1 gene from almond rootstock Nemaguard and transformed that into the Arabidopsis mutant (athkt1) to validate its functional role. The transgenic lines containing HKT1 from almond rootstock were tolerant to high salt concentrations, produced long and dense lateral roots, had low electrolyte leakage and maintained high relative water content. These observations suggest that transgenic plants coped well with increased salt concentration by maintain the integrity of the membranes. The HKT1 gene from almond rootstock was highly induced in Arabidopsis transgenic lines exposed to salt stress, confirming its functional validity. The functional characterization of candidate genes involved in salt tolerance will help in the development of tools for marker-assisted selection, which may become instrumental in screening almond germplasm to select novel sources for salt tolerance. The information generated will be useful to geneticists to decipher individual components of the salt tolerance mechanism, which then can be utilized by almond breeders to breed salt tolerant varieties.

Technical Abstract: Soil salinity affects plant growth and development, which directly impact yield. Plants deploy many mechanisms to cope with, or mitigate, salt stress. One of such mechanism is to control movement of ions from root to shoot by regulating the loading of Na+ in the transpiration stream. The high-affinity K+ transporter 1 (HKT1) is known to play a role in the removal of Na+ from the xylem and bring it back to the root. As almond is a salt-sensitive crop, the rootstock plays an important role in successful almond cultivation in salt-affected regions. We currently lack knowledge on the molecular mechanisms involved in salt tolerance of almond rootstocks. In this study, we complemented the Arabidopsis athkt1 knockout mutant with HKT1 ortholog (PpHKT1) from the almond rootstock ‘Nemaguard’. Arabidopsis trans-genic lines that were generated in athkt1 background with the constitutive promoter (PpHKT1OE2.2) and the native promoter (PpHKT1NP6) were subjected to different salt treatments. Both transgenic lines survived salt concentrations up to 120 mM NaCl, however, the mutant athkt1 died after 18 days under 120 mM NaCl. At 90 mM NaCl, the dry weight of athkt1 decreased significantly compared to the transgenic lines. Both transgenic lines showed significantly longer lateral roots compared to the athkt1 mutant at 80 mM NaCl treatment. The transgenic lines, PpHKT1OE2.2 and PpHKTNP6 had lower electrolyte leakage and higher relative water content compared to athkt1, suggesting that transgenic plants coped well with increased salt concentration by maintaining the integrity of the membranes. The expression analyses showed that PpHKT1 was induced in PpHKT1OE2.2 and PpHKTNP6 lines under salt treatment, which confirmed that both over-expression and native expression of PpHKT1 in the Arabidopsis mutant can complement salt tolerance function.