Physiology and proteomics research on the leaves of ancient Platycladus orientalis (L.) during winter

• Physiology and proteomic analysis of contrasting ancient and adult tree leaves during winter
• A number of protein species involved in various cellular pathways that are affected by low temperature stress
• A general reduction in photosynthesis and defense/stress-related proteins was observed in the leaves of ancient trees.
• Energy and carbohydrate metabolism protein species were abundant in the leaves of ancient trees.
• The analysis provides an understanding of the complex physiological changes that occur in ancient trees.

Ancient trees have an important value in humanities and history, and also have an important scientific value in the investigation of the decline and senescence mechanisms. Thus, we conducted an environmental stress study using ancient trees. To evaluate age-dependent changes in physiology and the leaf proteome, we assessed the low-temperature stress responses of 20 ± 5-, 500 ± 100- and 1200 ± 200-year-old Platycladus orientalis (L.) samples obtained outdoors during winter. Several physiological parameters were evaluated. Leaf proteomes were obtained using two-dimensional electrophoresis gels, and 77 protein spots were identified successfully using MALDI TOF/TOF MS/MS. The majority of the identified protein species were classified into functional categories including defense/stress-related, energy and carbohydrate metabolism, photosynthesis, and hormone-related functions. A general reduction in the abundance of protein species was observed as the age of the studied trees increased; reduction in photosynthesis and defense/stress-related categories were particularly apparent in the leaves of ancient trees. However, the number of protein species with functions in energy and carbohydrate metabolism increased with age. An increase in the abundance of lipid metabolism and hormone-related protein species was a primary characteristic of the leaves of ancient trees under low-temperature stress during winter. These results improve our understanding of the biochemical mechanisms of stress responses in ancient trees.Biological significanceLow temperature is the most common meteorological challenge in the study area. For evergreen plants, low-temperature stress has a great impact on the leaves of ancient P. orientalis. Thus, we conducted an environmental stress study using ancient trees. Recently, various studies were carried out in ancient trees. However, no information is available on the molecular mechanisms of defense to low-temperature stress in ancient trees. Therefore, this original study comprises the following differential proteomic analysis of ancient P. orientalis: (1) age-dependent changes in the physiology and leaf proteome are evaluated under low-temperature stress to (2) understand the differences in metabolic responses between ancient and adult trees under low-temperature stress during winter. This analysis will provide an understanding of the complex physiological changes that occur in ancient trees. The results suggest that certain identified proteins can be used as markers of low-temperature stress in ancient P. orientalis.

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