Thioredoxin and Redox Control within the New Concept of Oxidative Signaling

During the last decade, plant thioredoxins (TRX) h-type have been shown to be implicated in several new roles like the protection against the oxidative stress by their ability to reduce some antioxidant proteins as peroxiredoxins (PRX) or methionine-sulphoxide-reductases (MSR). However, the concept of the oxidative stress is changing and this fact raises the question of the TRX roles in this new context. In the January issue of Plant Physiology, we have presented two TRXsh from Pisum sativum differently involved in the control of the redox status. PsTRXh1 is an h-type TRX that acts by reducing classical antioxidant proteins. PsTRXh2 seems to be also involved in redox control, however it could act contrary to its counterpart h1. Both proteins may play antagonistic roles in pea in order to lead a better control of the redox status.Key Words: abiotic stress, oxidative signalling, thioredoxins, Pisum sativum, ROSHigh concentration of reactive oxygen species (ROS) in plant cells involve the activation of different antioxidant systems which reestablish the redox status leading to better physiological conditions. On the other hand, it has been well established that at certain levels, the ROS act as second messengers in signal transduction cascades in several processes in plant cells.¹ At the light of these events, it has been proposed the reevaluation of the concept of oxidative stress towards “oxidative signalling.”² This concept involves all the cellular mechanisms that let the plant cells sense and act in response to modified environmental conditions. Several cellular systems are involved in such role, and in these last years, plant TRXs have been shown to be involved in several number of metabolic pathways linked to the regulation of the redox imbalance,³ mainly for the case of the h-type cluster of the TRXs.^4–6In our last work,⁷ we have described two pea TRXs of the h-type cluster, PsTRXh1 and PsTRXh2 that are differentially, and even antagonistically, involved in the redoxregulation control, probably through their interaction with different target proteins. We proposed that PsTRXh1 might be involved in the control of the ROS levels in pea tissues due to its ability to interact with several antioxidant proteins in vivo. It is now very well known that some members of the TRX family reduce PRX,^8–10 an antioxidant enzyme involves in the direct deactivation of some oxidant agents or the MSR,^11,14,15 in charge of the recovery of the oxidized methionine, both in a very specific manner. Due to the increase of PsTRXh1 both at gene expression and protein levels in plant heterotrophic tissues in response to the H₂O₂ treatment, and because it is also capable of conferring resistance towards hydrogen peroxide when produced in a yeast trx1Δ trx2Δ strain,¹⁶ one function of this TRX member could be the reduction in vivo of some PRX and/or MSRA counterparts in Pea tissues in the context of the oxidative signalling.Interestingly, PsTRXh2 gene and its corresponding protein showed very different behaviours to that presented by its homologous h1, reinforcing the idea that some TRX isoforms in plants are capable of functional specificity in vivo. PsTRXh2 is expressed in all tissues assays, mainly in roots, but at an extremely low level compared with that of PsTRXh1. Its divergent functional behaviour was confirmed both in Pea plantlets and yeast. In fact, contrary to PsTRXh1, PsTRXh2 provides hypersensitivity in the yeast trx1Δ trx2Δ mutant. We explained the different behaviour by suggesting that PsTRXh2 might interact with some target(s) involved either directly or indirectly in hydrogen peroxide detoxification, either by compromising the target function in resistance to the ROS or by reinforcing the target function in producing sensitivity to H₂O₂. Most probably, PsTRXh1 and PsTRXh2 interact with very different partners, and the characterization of such targets may help in the deciphering of PsTRX isoforms. As short-term future experiments, using the TRX-specific two-hybrid system that was published recently,⁸ comparative efficiencies of PsTRX isoforms could be performed to reduce some putative target involved in H₂O₂ detoxification, including PsTRXh3 and PsTRXh4.¹⁷ Unravelling Pea TRX interactome should help in deciphering the function of each isoform.However, we have also considered the possibility that PsTRXh2 could interact with the same target that PsTRXh1, but producing an opposite effect. In the yeast context, the protein targeted by the heterologous plant TRXs responsible of this complementation is a type-II PRX.⁵ We think that PsTRXh2 could interact with this yeast PRX blocking it and producing the hypersensitivity. In fact, we have found a similar effect in other protein targeted in vitro by TRXs. In the (Fig. 1), we present the in vitro ability of PsTRXh1 and PsTRXh2 to reduce and activate the pea chloroplastic fructose-1,6-bisphosphatase (FBPase). In the presence of PsTRXh1, FBPase presents TRX-dependent activity but lower than that found when chloroplastic f and m1 isoforms are used.¹² On the opposite, PsTRXh2 presents no only FBPase activation capability but its presence induces the FBPase inhibition, as the enzymatic activity was lower than that exhibited by this enzyme without TRX.Open in a separate windowFigure 1TRX-dependent chloroplastic FBPase activity two-step procedure.¹³ , PsTRXf; ▴, PsTRXm1; •, PsTRXh1; , PsTRXh2. The negative control (no TRX) is represented by a symbol-less line. Numbers in each line represent maximum enzymatic velocity as OD₃₄₀/min.Considering all data, we think that the behaviour showed by both pea h-type TRXs is due to their interactions with several protein-targets, as the PRXs: when the ROS levels increase drastically, cells develop high redox imbalances or even undergo oxidative stress. In this situation, all antioxidant mechanisms must be activated, including the increase of PsTRXh1 expression and protein quantities, giving rise to a more efficient cell detoxification. Under nonimbalance conditions of the redox status, PsTRXh2 could act by interacting (activating or inhibiting) with some protein targets. However, the physiological target for PsTRXh2 are not yet described nor supposed. Our results suggest that its role in the redox control is by producing sensitivity to oxidant agents, maybe by allowing physiological ROS levels in cells.