TY - JOUR
T1 - A single residue deletion in the barley HKT1;5 P189 variant restores plasma membrane localisation but not Na+ conductance
AU - Wege, Stefanie
AU - Qiu, Jiaen
AU - Byrt, Caitlin
AU - Houston, Kelly
AU - Waugh, Robbie
AU - Gilliham, Matthew
AU - Hrmova, Maria
N1 - Funding -
SW, JQ, CB, and MG acknowledge funding from the Australian Research Council (CE140100008, DE160100804, and FT180100476). SW and CB thank the Grains Research and Development Corporation for their support. KH and RW acknowledge the Rural and Environment Science and Analytical Services Division, Scottish Government. MH credits the Huaiyin Normal University (China) for funding.
Copyright © 2018. Published by Elsevier B.V.
PY - 2021/10/1
Y1 - 2021/10/1
N2 - Leaf Na+ exclusion, mediated by plasma membrane-localised Class 1 High-affinity potassium (K+) Transporters (HKTs), is a key mechanism contributing to salinity tolerance of several major crop plants. We determined previously that the leucine to proline residue substitution at position 189 (L189P) in barley HvHKT1;5 disrupts its characteristic plasma membrane localisation and Na+ conductance. Here, we focus on a surprising observation that a single residue deletion of methionine at position 372 (M372del) within the conserved VMMYL motif in plant HKTs, restores plasma membrane localisation but not Na+ conductance in HvHKT1;5 P189. To clarify why the singular M372 deletion regains plasma membrane localisation, we built 3D models and defined α-helical assembly pathways of the P189 M372del mutant, and compared these findings to the wild-type protein, and the HvHKT1;5 L189 variant and its M372del mutant. We find that α-helical association and assembly pathways in HvHKT1;5 proteins fall in two contrasting categories. Inspections of structural flexibility through molecular dynamics simulations revealed that the conformational states of HvHKT1;5 P189 diverge from those of the L189 variant and M372del mutants. We propose that M372del in HvHKT1;5 P189 instigates structural rearrangements allowing routing to the plasma membrane, while the restoration of conductance would require further interventions. We integrate the microscopy, electrophysiology, and biocomputational data and discuss how a profound structural change in HvHKT1;5 P189 M372del impacts its α-helical protein association pathway and flexibility, and how these features underlie a delicate balance leading to restoring plasma membrane localisation but not Na+ conductance.
AB - Leaf Na+ exclusion, mediated by plasma membrane-localised Class 1 High-affinity potassium (K+) Transporters (HKTs), is a key mechanism contributing to salinity tolerance of several major crop plants. We determined previously that the leucine to proline residue substitution at position 189 (L189P) in barley HvHKT1;5 disrupts its characteristic plasma membrane localisation and Na+ conductance. Here, we focus on a surprising observation that a single residue deletion of methionine at position 372 (M372del) within the conserved VMMYL motif in plant HKTs, restores plasma membrane localisation but not Na+ conductance in HvHKT1;5 P189. To clarify why the singular M372 deletion regains plasma membrane localisation, we built 3D models and defined α-helical assembly pathways of the P189 M372del mutant, and compared these findings to the wild-type protein, and the HvHKT1;5 L189 variant and its M372del mutant. We find that α-helical association and assembly pathways in HvHKT1;5 proteins fall in two contrasting categories. Inspections of structural flexibility through molecular dynamics simulations revealed that the conformational states of HvHKT1;5 P189 diverge from those of the L189 variant and M372del mutants. We propose that M372del in HvHKT1;5 P189 instigates structural rearrangements allowing routing to the plasma membrane, while the restoration of conductance would require further interventions. We integrate the microscopy, electrophysiology, and biocomputational data and discuss how a profound structural change in HvHKT1;5 P189 M372del impacts its α-helical protein association pathway and flexibility, and how these features underlie a delicate balance leading to restoring plasma membrane localisation but not Na+ conductance.
KW - 3D protein modelling
KW - Confocal microscopy
KW - Molecular dynamics simulations
KW - Salinity
KW - Two-Electrode Voltage-Clamp
KW - α-Helical association and assembly pathways
UR - http://www.scopus.com/inward/record.url?scp=85109009501&partnerID=8YFLogxK
U2 - 10.1016/j.bbamem.2021.183669
DO - 10.1016/j.bbamem.2021.183669
M3 - Article
C2 - 34139196
VL - 1863
JO - BBA - Biomembranes
JF - BBA - Biomembranes
SN - 0005-2736
IS - 10
M1 - 183669
ER -