RESUMEN
The cyanide-resistant alternative oxidase (AOX) is a homodimeric protein whose activity can be regulated by the oxidation/reduction state and by alpha-keto acids. To further clarify the role of AOX in the skunk cabbage, Symplocarpus renifolius, we have performed expression and functional analyses of the encoding gene. Among the various tissues in the skunk cabbage, SrAOX transcripts were found to be specifically expressed in the thermogenic spadix. Moreover, our data demonstrate that the SrAOX protein exists as a non-covalently associated dimer in the thermogenic spadix, and is more sensitive to pyruvate than to other carboxylic acids. Our results suggest that the pyruvate-mediated modification of SrAOX activity plays a significant role in thermoregulation in the skunk cabbage.
Asunto(s)
Araceae/enzimología , Flores/enzimología , Oxidorreductasas/química , Oxidorreductasas/metabolismo , Ácido Pirúvico/farmacología , Temperatura , Secuencia de Aminoácidos , Araceae/efectos de los fármacos , Araceae/genética , Western Blotting , Ácidos Carboxílicos/farmacología , Respiración de la Célula/efectos de los fármacos , Diamida/farmacología , Dimerización , Ditiotreitol/farmacología , Flores/efectos de los fármacos , Flores/genética , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Ácidos Cetoglutáricos/farmacología , Mitocondrias/efectos de los fármacos , Mitocondrias/enzimología , Proteínas Mitocondriales , Datos de Secuencia Molecular , NAD/metabolismo , Oxidación-Reducción/efectos de los fármacos , Oxidorreductasas/genética , Proteínas de Plantas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Partículas Submitocóndricas/efectos de los fármacos , Partículas Submitocóndricas/enzimologíaRESUMEN
ent-Kaurene is a tetracyclic hydrocarbon precursor for gibberellins (GAs) in plants and fungi. To address whether fungal GA biosynthesis enzymes function in plants, we generated transgenic Arabidopsis plants overexpressing ent-kaurene synthase (GfCPS/KS) from a GA-producing fungus Gibberella fujikuroi. GfCPS/KS catalyzes a two-step reaction corresponding to ent-copalyl diphosphate synthase (CPS) and ent-kaurene synthase (KS) activities in plants. When GfCPS/KS was overexpressed and targeted to plastids, a range of GA-deficient phenotypes of the ga1-3 and ga2-1 mutants (defective in CPS and KS, respectively) were restored to wild type. Unexpectedly, the transgenic lines overproducing GfCPS/KS emitted the GA precursor ent-kaurene into the headspace besides its accumulation in the plant body. When co-cultivated with the ent-kaurene overproducers in a closed environment, the airborne ent-kaurene was able to fully complement the dwarf phenotype of ga1-3 and ga2-1 mutants, but not that of the ga3-1 mutant (defective in ent-kaurene oxidase). These results suggest that ent-kaurene may be efficiently metabolized into bioactive GAs in Arabidopsis when supplied as a volatile. We also provide evidence that ent-kaurene is released in the headspace of wild-type Chamaecyparis obtusa and Cryptomeria japonica plants, suggesting the occurrence of this hydrocarbon GA precursor as a volatile in nature.
