Potential involvement of N-terminal acetylation in the quantitative regulation of the ε subunit of chloroplast ATP synthase under drought stress.

In plants, modulation of photosynthetic energy conversion in varying environments is often accompanied by adjustment of the abundance of photosynthetic components. In wild watermelon (Citrullus lanatus L.), proteome analysis revealed that the ε subunit of chloroplast ATP synthase occurs as two distinct isoforms with largely-different isoelectric points, although encoded by a single gene. Mass spectrometry (MS) analysis of the ε isoforms indicated that the structural difference between the ε isoforms lies in the presence or absence of an acetyl group at the N-terminus. The protein level of the non-acetylated ε isoform preferentially decreased in drought, whereas the abundance of the acetylated ε isoform was unchanged. Moreover, metalloprotease activity that decomposed the ε subunit was detected in a leaf extract from drought-stressed plants. Furthermore, in vitro assay suggested that the non-acetylated ε subunit was more susceptible to degradation by metalloaminopeptidase. We propose a model in which quantitative regulation of the ε subunit involves N-terminal acetylation and stress-induced proteases.

Dynamic changes in the leaf proteome of a C3 xerophyte, Citrullus lanatus (wild watermelon), in response to water deficit.

Wild watermelon (Citrullus lanatus) is a xerophyte native to the Kalahari Desert, Africa. To better understand the molecular mechanisms of drought resistance in this plant, we examined changes in the proteome in response to water deficit. Wild watermelon leaves showed decreased transpiration and a concomitant increase in leaf temperature under water deficit conditions. Comparison of the proteome of stressed plants with that of unstressed plants by two-dimensional gel electrophoresis revealed that the intensity of 40 spots increased in response to the stress, and the intensity of 11 spots decreased. We positively identified 23 stress-induced and 6 stress-repressed proteins by mass spectrometry and database analyses. Interestingly, 15 out of the 23 up-regulated proteins (65% of annotated up-regulated proteins) were heat shock proteins (HSPs). Especially, 10 out of the 15 up-regulated HSPs belonged to the small heat shock protein (sHSP) family. Other stress-induced proteins included those related to antioxidative defense and carbohydrate metabolism. Fifteen distinct cDNA sequences encoding the sHSP were characterized from wild watermelon. Quantitative real-time PCR analysis of the representative sHSP genes revealed strong transcriptional up-regulation in the leaves under water deficit. Moreover, immunoblot analysis confirmed that protein abundance of sHSPs was massively increased under water deficit. Overall, these observations suggest that the defense response of wild watermelon may involve orchestrated regulation of a diverse array of functional proteins related to cellular defense and metabolism, of which HSPs may play a pivotal role on the protection of the plant under water deficit in the presence of strong light.

The long-term responses of the photosynthetic proton circuit to drought.

Proton motive force (pmf) across thylakoid membranes is not only for harnessing solar energy for photosynthetic CO(2) fixation, but also for triggering feedback regulation of photosystem II antenna. The mechanisms for balancing these two roles of the proton circuit under the long-term environmental stress, such as prolonged drought, have been poorly understood. In this study, we report on the response of wild watermelon thylakoid 'proton circuit' to drought stress using both in vivo spectroscopy and molecular analyses of the representative photosynthetic components. Although drought stress led to enhanced proton flux via a approximately 34% increase in cyclic electron flow around photosystem I (PS I), an observed approximately fivefold decrease in proton conductivity, g(H)(+), across thylakoid membranes suggested that decreased ATP synthase activity was the major factor for sustaining elevated q(E). Western blotting analyses revealed that ATP synthase content decreased significantly, suggesting that quantitative control of the complex plays a pivotal role in down-regulation of g(H)(+). The expression level of cytochrome b(6)f complex - another key control point in photosynthesis - also declined, probably to prevent excess-reduction of PS I electron acceptors. We conclude that plant acclimation to long-term environmental stress involves global changes in the photosynthetic proton circuit, in which ATP synthase represents the key control point for regulating the relationship between electron transfer and pmf.