Vitamin B6 （vitB6） serves as an essential cofactor for more than 140 enzymes. Pyridoxal 5＇-phosphate （PLP）, active cofactor form of vitB6, can be photolytically destroyed by trace amounts of ultraviolet-B （UV-B...Vitamin B6 （vitB6） serves as an essential cofactor for more than 140 enzymes. Pyridoxal 5＇-phosphate （PLP）, active cofactor form of vitB6, can be photolytically destroyed by trace amounts of ultraviolet-B （UV-B）. How sun-exposed organisms cope with PLP photosensitivity and modulate vitB6 homeostasis is currently unknown. We previously reported on two Arabidopsis mutants, rusl and rus2, that are hypersensitive to trace amounts of UV-B light. We performed mutagenesis screens for second-site suppressors of the rus mutant phenotype and identified mutations in the ASPARTATE AMINOTRANSFERASE2 （ASP2） gene. ASP2 encodes for cytosolic aspartate aminotransferase （AAT）, a PLP-dependent enzyme that plays a key role in carbon and nitrogen metabolism. Genetic analyses have shown that specific amino acid substitutions in ASP2 override the phenotypes of rusl and rus2 single mutants as well as rusl rus2 double mutant. These substitutions, all shown to reside at specific positions in the PLP-binding pocket, resulted in no PLP binding. Additional asp2 mutants that abolish AAT enzymatic activity, but which alter amino acids outside of the PLP-binding pocket, fail to suppress the rus phenotype. Furthermore, exogenously adding vitB6 in growth media can rescue both rusl and rus2. Our data suggest that AAT plays a role in vitB6 homeostasis in Arabidopsis.展开更多
文摘Vitamin B6 （vitB6） serves as an essential cofactor for more than 140 enzymes. Pyridoxal 5＇-phosphate （PLP）, active cofactor form of vitB6, can be photolytically destroyed by trace amounts of ultraviolet-B （UV-B）. How sun-exposed organisms cope with PLP photosensitivity and modulate vitB6 homeostasis is currently unknown. We previously reported on two Arabidopsis mutants, rusl and rus2, that are hypersensitive to trace amounts of UV-B light. We performed mutagenesis screens for second-site suppressors of the rus mutant phenotype and identified mutations in the ASPARTATE AMINOTRANSFERASE2 （ASP2） gene. ASP2 encodes for cytosolic aspartate aminotransferase （AAT）, a PLP-dependent enzyme that plays a key role in carbon and nitrogen metabolism. Genetic analyses have shown that specific amino acid substitutions in ASP2 override the phenotypes of rusl and rus2 single mutants as well as rusl rus2 double mutant. These substitutions, all shown to reside at specific positions in the PLP-binding pocket, resulted in no PLP binding. Additional asp2 mutants that abolish AAT enzymatic activity, but which alter amino acids outside of the PLP-binding pocket, fail to suppress the rus phenotype. Furthermore, exogenously adding vitB6 in growth media can rescue both rusl and rus2. Our data suggest that AAT plays a role in vitB6 homeostasis in Arabidopsis.
