A pivotal role of phosphatidylinositol 3-kinase in delaying of methyl jasmonate-induced leaf senescence

ABSTRACT

Phosphatidylinositol 3-kinase (PI3K) and its product PI3P are involved in plant development and stress responses. Our recent report has suggested that down-regulation of PI3K activity accelerated leaf senescence induced by methyl jasmonate (MeJA) and suppressed the activation of vacuolar H⁺-ATPase (V-ATPase). In vitro and in vivo experiment revealed that PI3K interact with VHA-B2. The inhibition of V-ATPase activity suppressed the vacuolar acidification and enhanced the stomatal opening, thereby accelerating MeJA-induced leaf senescence. It was shown that there is close relationship between PI3K and V-ATPase. However, the factor which initiates the PI3K-V-ATPase pathway needs further improvement, and the domain of VHA-B that binds to PI3K is still not clear enough. By using the Arabidopsis and MeJA as the research model, studies have been performed to investigate the upstream regulation of PI3K and downstream function of PI3K-V-ATPase pathway in the plant senescence.

Emerging evidence suggests that water maintenance is important for the delay of leaf senescence.¹ As we know, water control is mostly dependent on stomatal movement. Recent report has reported that vacuolar acidification has a key role in ABA-induced stomatal closure.² In plants, vacuolar H⁺-ATPase (V-ATPase) is a key proton pump localized in tonoplasts. The major mechanism of V-ATPase regulation is the reversible assembly/disassembly of the V0 and V1 sectors.³ In renal epithelial cells induced with glucose, phosphoinositide 3-kinase type I (PI3K) promotes the assembly and translocation of V-ATPase.⁴ PI3K activated by influenza A virus also interacts with the subunit E of the V1 domain of V-ATPase.⁵ These findings prompt us to further investigate the signaling mechanism of V-ATPase and the important role of PI3K in MeJA-induced leaf senescence.

Accumulating experimental evidence support that water loss of senescing leaves is faster than that of non-senescing leaves.¹ Our current work has shown that stomatal opening was enhanced with prolonged MeJA treatment and vacuolar lumen was alkalized. The enhanced stomatal aperture was also found in low-acidified vacuolar lumen in MeJA-induced leaf senescence. Then, as the main proton pump localized in tonoplast, V-ATPase activity and gene expression of subunits were investigated. Additionally, stomatal opening was enhanced compared with that under control conditions, when treated senescent leaves induced by MeJA with the V-ATPase inhibitor Bafilomycin A1 (BFA1) to accelerate vacuolar alkalization.⁶ However, another type of proton pump, vacuolar H⁺-pyrophosphatases (V-PPases) cannot be ignored. V-PPases are single-subunit homodimers that also generate proton gradients in endomembrane compartments by using pyrophosphate (PPi) other than ATP.⁷ Previous study reported that over-expression of V-PPase induced stomatal closure and reduced the number of vacuoles in guard cells.⁸ Therefore, the effect of V-PPase on vacuolar acidification should be investigated in MeJA-induced leaf senescence too.

The former study revealed that PI3K could bind with V-ATPase subunit in animals. Our data determined that PI3K also bind with V-ATPase subunit in plant.⁶ Additionally, BiFC experiment was further conducted using onion epidermal cells to prove the interaction of PI3K with the V-ATPase subunit B2 on tonoplast. Tonoplast localization was achieved by fusing the coding region of the mCherry to the C-terminus of γ-TIP (VAC-mCherry), an aquaporin of the vacuolar membrane.⁹ Clear YFP fluorescence signal was observed on the tonoplast of onion epidermal cells. However, we cannot exclude the other possible subcellular localization. PI3K type III plays a vital role in the trafficking of proteins to and from vacuoles.¹⁰ Interactions between VHA-B2 and PI3K are possibly promoted the translocation of VHA-B2 from the TNG to the vacuole and thus improved the assembly of V-ATPase, resulting in increased V-ATPase activity. These aspects should be investigated in the future.

Our data demonstrated that vacuolar acidification was regulated by the PI3K-V-ATPase pathway in the MeJA-induced leaf senescence. The inhibition of PI3K or V-ATPase activity leads to the early senescence.⁶ However, PI3K also exhibited diverse functions in plant development and stress response, such as actin filament reorganization, plasma membrane endocytosis, and NADPH oxidase activation.^11-13 Hence, it may be involved in the regulation of leaf senescence.

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Funding

This research is supported by the Program for Changjiang Scholars and Innovative Research Team in University (IRT0829), the Key Program of NSFC-Guangdong Joint Funds of China (U0931005), the National High Technology Research and Development Program of China (863 Program) (2007AA10Z204).