Expression, in vivo localization and phylogenetic analysis of a pyridoxine 5'-phosphate oxidase in Arabidopsis thaliana.
Pyridoxal phosphate (PLP), a vitamin B(6) vitamer, is an essential cofactor for numerous enzymes. Pyridoxine/pyridoxamine phosphate oxidase (PPOX) catalyzes the synthesis of pyridoxal phosphate from pyridoxine phosphate (PNP) and/or pyridoxamine phosphate (PMP). The At5g49970 locus in Arabidopsis thaliana encodes an AtPPOX, a PNP/PMP oxidase. The expression of the AtPPOX gene varied in different tissues of Arabidopsis examined, being up-regulated by light, heat shock, ABA, and ethylene treatments, and down-regulated by exposure to drought and NaCl. Monoclonal antibodies raised against two different domains of AtPPOX recognized different sizes of AtPPOX, suggesting that AtPPOX proteins are produced as splice variants of the AtPPOX gene in Arabidopsis. Phylogenetic analysis of AtPPOX across all domains of life demonstrated that plant AtPPOX homologs have an additional Yjef_N domain preceding the Pyridox_Oxidase domain at the C-terminal end of the protein, while AtPPOX homologs from bacteria, fungi and animals have only Pyridox_Oxidase domain. The presence of the Yjef_N domain in plant AtPPOX homologs suggests that acquisition of this domain, and its fusion with the pyridox_oxidase domain began with the endosymbiotic acquisition of the chloroplast. Bioinformatic analysis suggested that AtPPOX is localized in chloroplast, but the monoclonal antibody could not be used for subcellular localization of this protein. A GFP-AtPPOX fusion construct introduced into the Arabidopsis protoplast confirmed localization of AtPPOX into the chloroplast. An RNAi mutant of AtPPOX showed sensitivity to high light suggesting a role for PPOX in resistance to photooxidative damage, and alteration in root growth in the presence of sucrose suggests a role for PPOX in root development.
Department of Biological Sciences, 101 Rouse Life Sciences Building, Auburn University, AL 36849, USA.
Yuying
Y
Sang
Robert D
RD
Locy
Leslie R
LR
Goertzen
Aaron M
AM
Rashotte
Ying
Y
Si
Kwonkyoo
K
Kang
Narendra K
NK
Singh
D000911:Antibodies, Monoclonal; D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics; D029681:Arabidopsis Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D001419:Bacteria / Q000235:genetics; D002736:Chloroplasts / Q000378:metabolism; D005658:Fungi / Q000235:genetics; D015870:Gene Expression; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D008027:Light; D009154:Mutation; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010802:Phylogeny; D018517:Plant Roots / Q000254:growth & development; D011523:Protoplasts; D011734:Pyridoxaminephosphate Oxidase / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D034741:RNA, Small Interfering; D013559:Symbiosis
D000911:Antibodies, Monoclonal; D029681:Arabidopsis Proteins; D034741:RNA, Small Interfering; D011734:Pyridoxaminephosphate Oxidase