Cloning and functional expression of the mitochondrial alternative oxidase of Aspergillus fumigatus and its induction by oxidative stress.
Aspergillus fumigatus possesses a branched mitochondrial electron transport chain, with both cyanide-sensitive and -insensitive oxygen-consumption activities. Mitochondrial reactive oxygen species mediate signaling for alternative oxidase (AOX) expression. A 1173 bp-long Afaox gene encoding a 40 kDa protein has been cloned and identified. Recombinant constructs containing the Afaox ORF were transformed into Escherichia coli and Saccharomyces cerevisiae for heterologous expression. In A. fumigatus, AOX activity and mRNA expression were both induced with menadione or paraquat, suggesting an important role of AOX under oxidative stress. Therefore, positive transformants showed a cyanide-resistant and salicylhydroxamic acid-sensitive respiration, whereas in control cells the oxygen uptake was completely inhibited after KCN addition.
Magnani
T
T
Dep Análises Clínicas, Toxicológicas e Bromatológicas, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, SP, Brazil.
Soriani
F M
FM
Martins
V P
VP
Nascimento
A M
AM
Tudella
V G
VG
Curti
C
C
Uyemura
S A
SA
D000595:Amino Acid Sequence; D001232:Aspergillus fumigatus / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D015153:Blotting, Western; D003001:Cloning, Molecular; D003486:Cyanides / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal; D008928:Mitochondria / Q000201:enzymology / Q000378:metabolism; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D010088:Oxidoreductases / Q000235:genetics* / Q000378:metabolism; D010101:Oxygen Consumption / Q000187:drug effects; D010269:Paraquat / Q000494:pharmacology; D010940:Plant Proteins; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D012457:Salicylamides / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D024483:Vitamin K 3 / Q000494:pharmacology; D000099188:Alternative Oxidase
D003486:Cyanides; D005656:Fungal Proteins; D024101:Mitochondrial Proteins; D010088:Oxidoreductases; D010269:Paraquat; D010940:Plant Proteins; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D012457:Salicylamides; D024483:Vitamin K 3; D000099188:Alternative Oxidase; C005703:salicylhydroxamic acid