Molecular characterization and expression analysis of the Na+/H+ exchanger gene family in Medicago truncatula

Authors: Sandhu, Devinder; Pudussery, Manju; KAUNDAL, RAKESH - Utah State University; Suarez, Donald; KAUNDAL, AMITA - University Of California; SEKHON, RAJANDEEP - Clemson University

Submitted to: Functional and Integrative Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/8/2017
Publication Date: 12/26/2017
Citation: Sandhu, D., Pudussery, M.V., Kaundal, R., Suarez, D.L., Kaundal, A., Sekhon, R.S. 2018. Molecular characterization and expression analysis of the Na+/H+ exchanger gene family in Medicago truncatula. Functional and Integrative Genomics. 18(2):141-153. https://doi.org/10.1007/s10142-017-0581-9.
DOI: https://doi.org/10.1007/s10142-017-0581-9

Interpretive Summary: Medicago truncatula is an ideal model legume. As M. truncatula DNA is completely sequenced, it can be used to study genetic networks involved in salt tolerance in alfalfa. Plants develop various mechanisms and strategies to cope with salinity. One important mechanism is keeping the cytosolic Na+ concentration low by sequestering Na+ in vacuoles, a process facilitated by Na+/H+ exchangers (NHX). There are eight NHX genes (NHX1 through NHX8) identified and characterized in Arabidopsis. We used Arabidopsis genes and performed bioinformatic analysis to identify six corresponding Medicago truncatula genes (MtNHX1, MtNHX2 MtNHX3 MtNHX4 MtNHX6 and MtNHX7). There were no corresponding genes identified for AtNHX5 and AtNHX8. Similar structure, organization and evolutionary relationship of NHX proteins in Arabidopsis and M. truncatula suggest similar functional roles of these proteins in two species. In salinity stress experiments, M. truncatula exhibited ~20% reduction in biomass. Sodium contents increased by 178% and 75% in leaves and roots, respectively and Cl- contents increased by 152% and 162%, respectively, in the salinity treatment. Na+ exclusion may be responsible for the relatively smaller increase in Na+ concentration as compared to Cl- under salt stress. As seen for the NHX genes in other plants, different MtNHXs were expressed in all the organs tested, however, the expression levels varied for different NHX genes. Expression analysis results suggest that in M. truncatula sequestering Na+ into vacuoles may not be the principle component trait of the salt tolerance mechanism and other component traits may be pivotal. This analysis can be used to compare similarities and differences in the salt tolerance mechanism in M. truncatula and alfalfa. The information generated will be useful to geneticists to characterize genes critical for salt tolerance in alfalfa and to use them to breed salt tolerant varieties.

Technical Abstract: One important mechanism plants use to cope with salinity is keeping the cytosolic Na+ concentration low by sequestering Na+ in vacuoles, a process facilitated by Na+/H+ exchangers (NHX). There are eight NHX genes (NHX1 through NHX8) identified and characterized in Arabidopsis. Bioinformatic analysis of the known Arabidopsis genes enabled us to identify six corresponding Medicago truncatula genes (MtNHX1, MtNHX2 MtNHX3 MtNHX4 MtNHX6 and MtNHX7). Twelve transmembrane domains and an amiloride binding site were conserved in five out of six MtNHX proteins. Phylogenetic analysis involving A. thaliana, Glycine max, Phaseolus vulgaris and M. truncatula revealed that each individual MtNHX class (Class I: MtNHX1 through 4; Class II: MtNHX6; Class III: MtNHX7) falls under a separate clade. In salinity-stress experiment, M. truncatula exhibited ~20% reduction in biomass. In the salinity treatment, sodium contents increased by 178% and 75% in leaves and roots, respectively and Cl- contents increased by 152% and 162%, respectively. Na+ exclusion may be responsible for the relatively smaller increase in Na+ concentration in roots under salt stress as compared to Cl-. Decline in tissue K+ concentration under salinity was not surprising as some antiporters play important role in transporting both Na+ and K+. MtNHX1, MtNHX6 and MtNHX7 display high expression in roots and leaves. MtNHX3, MtNHX6, MtNHX7 were induced in roots under salinity stress. Expression analyses results indicate that in M. truncatula sequestering Na+ into vacuoles may not be the principle component trait of the salt tolerance mechanism and other component traits may be pivotal.