title: Alkaline alpha-galactosidase degrades thylakoid membranes in the chloroplast during leaf senescence in rice.
abstract: Here, we studied the functional role of a chloroplast alkaline alpha-galactosidase (OsAkalphaGal) in the breakdown of thylakoid membranes during rice (Oryza sativa) leaf senescence. We assayed the enzyme activity of recombinant OsAkalphaGal with different natural substrates and examined the effect of ectopic OsAkalphaGal expression in rice plants. Recombinant OsAkalphaGal showed at least a two-fold greater substrate-binding affinity and a 10-fold greater turnover rate to galactolipid digalactosyl diacylglycerol than the raffinose family of oligosaccharides (verbascose, stachyose, raffinose) and melibiose. The OsAkalphaGal null mutant (osakalphagal) displayed a delayed leaf senescence phenotype. OsAkalphaGal complementation in osakalphagal recovered OsAkalphaGal expression and showed a senescence phenotype similar to that of wild-type plants. Transgenic plants overexpressing OsAkalphaGal (UbiP-OsAkalphaGal) exhibited retarded plant growth and development, and showed a pale-green phenotype coupled with a reduced chlorophyll content to 42% in newly unfolded leaves. UbiP-OsAkalphaGal leaves also showed a 29-fold increase in alkaline alpha-galactosidase activity compared with wild-type leaves. An ultrastructural study of Ubi-OsAkalphaGal chloroplasts in newly unfolded leaves revealed abnormal grana organization. Our findings strongly suggest that OsAkalphaGal is a thylakoid membrane-degrading enzyme involved in the degradation of digalactosyl diacylglycerol during rice leaf senescence.
authors.affiliation: Department of Life Sciences and Institute of Biodiversity, National Cheng-Kung University, No. 1, University Road, Tainan 700, Taiwan.
authors.forename: Ruey-Hua
authors.initials: RH
authors.lastname: Lee
authors.affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Nankang, Taipei, Taiwan.
authors.forename: Jen-Hung
authors.initials: JH
authors.lastname: Hsu
authors.affiliation: Department of Life Sciences and Institute of Biodiversity, National Cheng-Kung University, No. 1, University Road, Tainan 700, Taiwan.
authors.forename: Hao-Jen
authors.initials: HJ
authors.lastname: Huang
authors.affiliation: Chiayi Agricultural Experiment Station, TARI, Chiayi, Taiwan.
authors.forename: Shu-Fang
authors.initials: SF
authors.lastname: Lo
authors.affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Nankang, Taipei, Taiwan.
authors.forename: Shu-Chen
authors.initials: SC
authors.lastname: Grace Chen
mesh_terms: D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D038983:Galactolipids / Q000378:metabolism; D017353:Gene Deletion; D017343:Genes, Plant; D005816:Genetic Complementation Test; D007700:Kinetics; D046529:Microscopy, Electron, Transmission; D012275:Oryza / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000648:ultrastructure; D010641:Phenotype; D018515:Plant Leaves / Q000201:enzymology / Q000254:growth & development / Q000648:ultrastructure; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D020524:Thylakoids / Q000201:enzymology* / Q000648:ultrastructure; D000519:alpha-Galactosidase / Q000235:genetics / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D018744:DNA, Plant; D038983:Galactolipids; D010940:Plant Proteins; D011994:Recombinant Proteins; C007388:digalactosyldiacylglycerol; D000519:alpha-Galactosidase
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