Abstract
As an important primary metabolite, malate plays a key role in regulating osmotic pressure, pH homeostasis, stress tolerance, and fruit quality of apple. The R2R3-MYB transcription factor (TF) MdMYB73 was identified as a protein that plays a critical role in determining malate accumulation and vacuolar acidification by directly regulating the transcription of aluminum-activated malate transporter 9 (MdALMT9), vacuolar ATPase subunit A (MdVHA-A), and vacuolar pyrophosphatase 1 (MdVHP1) in apple. In addition, the bHLH TF MdCIbHLH1 interacts with MdMYB73 and enhances the transcriptional activity of MdMYB73. Our previous studies demonstrated that the BTB-BACK-TAZ domain protein MdBT2 can degrade MdCIbHLH1 to influence malate accumulation and vacuolar acidification. However, the potential upstream regulators of MdMYB73 are currently unknown. In this study, we found that MdBT2 directly interacts with and degrades MdMYB73 through the ubiquitin/26S proteasome pathway to regulate malate accumulation and vacuolar acidification. A series of functional assays with apple calli and fruit showed that MdBT2 controls malate accumulation and vacuolar acidification in an MdMYB73-dependent manner. Overall, our findings shed light on the mechanism by which the BTB-BACK-TAZ domain protein MdBT2 regulates malate accumulation and vacuolar acidification by targeting MdMYB73 and MdCIbHLH1 for ubiquitination in apple. This information may help guide traditional breeding programs and fruit tree molecular breeding, and lead to improvements in fruit quality and stress tolerance.
Highlights
Malate, a major intermediate in several metabolic processes, is present in most plant species and plays critical roles in regulating osmotic pressure, pH homeostasis, nutrient absorption, and stress resistance[1,2]
The full-length cDNA and five truncated mutants of the MdBT2 gene were independently inserted into the pGBT9 vector as the bait vectors, while the fulllength cDNA of MdMYB73 was inserted into the pGAD424 vector as the prey vector
Malate is a pivotal metabolite that is involved in regulating osmotic pressure, pH homeostasis, and stress resistance, and it significantly influences fruit quality[2,7]
Summary
A major intermediate in several metabolic processes, is present in most plant species and plays critical roles in regulating osmotic pressure, pH homeostasis, nutrient absorption, and stress resistance[1,2]. Malate transport involves vacuolar proton pumps, and malate transporters play key roles in malate transport from the cytosol into the vacuole, driving vacuolar acidification[2]. Vacuolar proton pumps contain vacuolar H+-ATPase (VATPase) and H+-pyrophosphatase (V-PPase)[12]. Zhang et al Horticulture Research (2020)7:151 has a complex structure that includes two functional parts: the peripheral V1 sector and the membrane-embedded V0 sector[13,14]. V1 has eight different subunits, with A and B subunits critical for the catalytic activity, and C–H subunits linked to hydrophobic and membrane-embedded V0, as is critical for ATP hydrolysis[12,15]. V-ATPase and V-PPase are both necessary for forming and maintaining the electrochemical potential gradient on the tonoplast[2,19], and for supplying energy to other transport proteins, such as the malate transporters tDT, Ma1, and Ma10 V-ATPase and V-PPase are both necessary for forming and maintaining the electrochemical potential gradient on the tonoplast[2,19], and for supplying energy to other transport proteins, such as the malate transporters tDT, Ma1, and Ma10 (refs. 7,12,20–22)
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