A bioassay-guided fractionation system to identify endogenous small molecules that activate plasma membrane H+-ATPase activity in Arabidopsis

Abstract

Endogenous unsaturated fatty acids identified from salt-treated Arabidopsis by a bioassay-guided fractionation method activate plasma membrane H+-ATPase by binding to its C-terminus.

Root-zone-specific sensitivity of K+-and Ca2+-permeable channels to H2O2 determines ion homeostasis in salinized diploid and hexaploid Ipomoea trifida

“…PM H + -ATPase activity, which is usually reflected by the kinetics and magnitude of H + efflux in plant cells (Bose et al , 2013; Yu et al , 2016; Han et al , 2017), is essential for the maintenance of membrane potential across the PM, prevents K + loss through DA-KORCs, and contributes to the retention of high cytosolic K + levels under saline conditions (Sun et al , 2009 a ; Anschütz et al , 2014; Yu et al ., 2016, 2018). The intrinsically high H + -ATPase activity in specific root zones contributes to the low salt-induced K + efflux in barley (Shabala et al , 2016 b ).…”

Elucidating the molecular mechanisms mediating plant salt‐stress responses

“…Melatonin accumulation in plant cells confers salt tolerance by reducing ROS and delaying leaf senescence (Liang et al ., ). Free unsaturated fatty acids stimulate the activity of the PM H + ‐ATPase under salt stress (Han et al ., ). Currently, it is not understood in detail how these small molecules enhance stress tolerance in plants.…”

“…Under salt stress, both 14‐3‐3 and GI are degraded by the 26S proteasomal pathway (Kim et al ., ; Tan et al ., ), and PKS5 activity is repressed to release PM H + ‐ATPase activity (Yang et al ., ). Under salt stress, free unsaturated fatty acids activate the PM H + ‐ATPase by directly binding its C‐terminus in Arabidopsis root cells (Han et al ., ), and the accumulation of fatty acids is required for plant salt resistance (Zhang et al ., ).…”

An improved protein lipid overlay assay for studying lipid–protein interactions

“…By coupling ATP hydrolysis and proton transport, PM H + -ATPase establishes a membrane potential for secondary transporters to transporting many ions and nutrients [10]. In our previous study, we found that endogenous oleic acid (C18:1), linoleic acid (C18:2) and linolenic acid (C18:3) can stimulate PM H + -ATPase activity in Arabidopsis, and interaction assays confirmed that all three lipids bound to the C-terminus of the PM H + -ATPase AHA2 protein [13]. Fatty acids were dissolved in 1:1 dichloromethane:MeOH and the MPLO assay was performed.…”

“…The cloning, expression, and purification of AHA2 central loop, AHA2 C-terminus, AtROP6, CKL2, and MPK6 were performed similarly as previously reported [12,13]. Briefly, the coding sequences of AHA2 central loop, AHA2 C-terminus, CKL2, and MPK6 from Arabidopsis were amplified by RT-PCR with the primer of L-Bf/L-Er for AHA2 central loop, C-Bf/C-Er for AHA2 C-terminus, CKL2-Bf/CKL2-Er for CKL2, and MPK6-Bf/MPK6-Hr for MPK6, respectively.…”