title: Molecular and biochemical characterization of a cyanogenic β-glucosidase in the inner bark tissues of rubber tree (Hevea brasiliensis Muell. Arg.).
abstract: Tapping causes the loss of large amounts of latex from laticifers and subsequently enhances latex regeneration, a high carbon- and nitrogen-cost activity in rubber tree. It is suggested that a 67 kDa protein associated with protein-storing cells in the inner bark tissues of rubber tree plays an important role in meeting the nitrogen demand for latex regeneration. Here, the 67 kDa protein was further characterized by a combination of cell biological, molecular biological and biochemical techniques. Immunogold labeling showed that the 67 kDa protein was specifically localized in the central vacuole of protein-storing cells. A full-length cDNA, referred to as HbVSP1, was cloned. The HbVSP1 contained a 1584 bp open reading frame encoding a protein of 527 amino acids. The putative protein HbVSP1 shared high identity with the P66 protein from rubber tree and proteins of the linamarase, and bg1A from cassava (Manihot esculenta). HbVSP1 contained the active site sequences of β-glucosidase, TFNEP and I/VTENG. In vitro analysis showed that the 67 kDa protein exhibited the activity of both β-glucosidase and linamarase and was thus characterized as a cyanogenic β-glucosidase. Proteins immuno-related to the 67 kDa protein were present in leaves and lutoids of laticifers. Tapping down-regulated the expression of HbVSP1, but up-regulated the expression of genes encoding the key enzymes for rubber biosynthesis, while the effect of resting from tapping was the reverse. Taken together, the results suggest that the 67 kDa protein is a vacuole-localized cyanogenic β-glucosidase encoded by HbVSP1 and may have a role in nitrogen storage in inner bark tissues of trunk during the leafless periods when rubber tree is rested from tapping.
authors.affiliation: Ministry of Agriculture Key Laboratory of Biology and Genetic Resources of Rubber Tree, Rubber Research Institute, Chinese Academy of Tropical Agricultural Sciences, Danzhou, Hainan 571737, PR China. wmtian@163.com
authors.forename: Wei-Min
authors.initials: WM
authors.lastname: Tian
authors.forename: Hua
authors.initials: H
authors.lastname: Zhang
authors.forename: Shu-Guang
authors.initials: SG
authors.lastname: Yang
authors.forename: Min-Jing
authors.initials: MJ
authors.lastname: Shi
authors.forename: Xu-Chu
authors.initials: XC
authors.lastname: Wang
authors.forename: Long-Jun
authors.initials: LJ
authors.lastname: Dai
authors.forename: Yue-Yi
authors.initials: YY
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D018506:Gene Expression Regulation, Plant; D028482:Hevea / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D024301:Plant Bark / Q000201:enzymology; D012408:Rubber / Q000378:metabolism*; D001617:beta-Glucosidase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
chemical_list: D012408:Rubber; C021514:cyanogenic beta-glucosidase; D001617:beta-Glucosidase