No abstract

PAO specifically oxidizes substrates that have both primary and secondary amino groups. The complex with MDL72527 shows that the primary amino groups are essential for the proper alignment of the substrate with respect to the flavin. Conservation of an N-terminal sequence motif indicates that PAO is member of a novel family of flavoenzymes. Among these, monoamine oxidase displays significant sequence homology with PAO, suggesting a similar overall folding topology.

Polyamine oxidases (PAOs) are FAD-dependent enzymes involved in polyamine catabolism. All so far characterized PAOs from monocotyledonous plants, such as the apoplastic maize PAO, oxidize spermine (Spm) and spermidine (Spd) to produce 1,3-diaminopropane, H(2)O(2), and an aminoaldehyde, and are thus considered to be involved in a terminal catabolic pathway. Mammalian PAOs oxidize Spm or Spd (and/or their acetyl derivatives) differently from monocotyledonous PAOs, producing Spd or putrescine, respectively, in addition to H(2)O(2) and an aminoaldehyde, and are therefore involved in a polyamine back-conversion pathway. In Arabidopsis thaliana, five PAOs (AtPAO1-AtPAO5) are present with cytosolic or peroxisomal localization and three of them (the peroxisomal AtPAO2, AtPAO3, and AtPAO4) form a distinct PAO subfamily. Here, a comparative study of the catalytic properties of recombinant AtPAO1, AtPAO2, AtPAO3, and AtPAO4 is presented, which shows that all four enzymes strongly resemble their mammalian counterparts, being able to oxidize the common polyamines Spd and/or Spm through a polyamine back-conversion pathway. The existence of this pathway in Arabidopsis plants is also evidenced in vivo. These enzymes are also able to oxidize the naturally occurring uncommon polyamines norspermine and thermospermine, the latter being involved in important plant developmental processes. Furthermore, data herein reveal some important differences in substrate specificity among the various AtPAOs, which suggest functional diversity inside the AtPAO gene family. These results represent a new starting point for further understanding of the physiological role(s) of the polyamine catabolic pathways in plants.

Wounding chickpea (Cicer arietinum) internodes or cotyledons resulted in an increase in the steady-state level of copper amine oxidase (CuAO) expression both locally and systemically. Dissection of the molecular mechanisms controlling CuAO expression indicated that jasmonic acid worked as a potent inducer of the basal and wound-inducible CuAO expression, whereas salicylic acid and abscisic acid caused a strong reduction of the wound-induced CuAO expression, without having any effect on the basal levels. Epicotyl treatment with the CuAO mechanism-based inhibitor 2-bromoethylamine decreased hydrogen peroxide (H 2 O 2 ) levels in all the internodes, as evidenced in vivo by 3,3Ј-diaminobenzidine oxidation. Moreover, inhibitor pretreatment of wounded epicotyls resulted in a lower accumulation of H 2 O 2 both at the wound site and in distal organs. In vivo CuAO inhibition by 2-bromoethylamine after inoculation of resistant chickpea cv Sultano with Ascochyta rabiei resulted in the development of extended necrotic lesions, with extensive cell damage occurring in sclerenchyma and cortical parenchyma tissues. These results, besides stressing the fine-tuning by key signaling molecules in wound-induced CuAO regulation, demonstrate that local and systemic CuAO induction is essential for H 2 O 2 production in response to wounding and indicate the relevance of these enzymes in protection against pathogens.Plant defense responses are accomplished by the deployment of a complex array of events that are differentially modulated depending on the incoming stress (Maleck and Dietrich, 1999). Wounding different plant organs or interaction with pathogens induce local and systemic accumulation of defenserelated proteins (Hammond-Kosack and Jones, 1996;Ryals et al., 1996;Ryan, 2000). The study of signaling events inducing local and systemic responses led to the discovery of systemin, jasmonates, ethylene, salicylic acid (SA), and abscisic acid (ABA) as signal molecules (Peñ a-Cortés et al., 1989; Farmer and Ryan, 1990; Pearce et al., 1991;Xu et al., 1994; O'Donnell et al., 1996;Schweizer et al., 1998;van Loon et al., 1998; Knoester et al., 1999).The existence of multiple defense strategies and complex signaling networks leads to an enhanced defense capacity of the plants. The signal transduction pathways of wounding and pathogen attack may be common, different, or exclusive, depending on the biological system, but likewise the establishment of defense mechanisms requires the presence or accumulation of hydrogen peroxide (H 2 O 2 ; Sutherland, 1991; Mehdy, 1994; Hammond-Kosack et al., 1996). In particular, H 2 O 2 behaves as a direct cytotoxic compound against pathogens and as a second messenger in the activation of defense genes (Lamb and Dixon, 1997). Moreover, this compound is involved in systemic acquired resistance and acts synergistically with NO in the induction of hypersensitive cell death (Delledonne et al., 1998). As a cosubstrate of the peroxidases, H 2 O 2 has been implicated in the oxidative cross-linking of apoplastic st...