Mitochondrial transport in proline catabolism in plants: the existence of two separate translocators in mitochondria isolated from durum wheat seedlings

Abstract

Abiotic stresses, such as high salinity or drought, can cause proline accumulation in plants. Such an accumulation involves proline transport into mitochondria where proline catabolism occurs. By using durum wheat seedlings as a plant model system, we investigated how proline enters isolated coupled mitochondria. The occurrence of two separate translocators for proline, namely a carrier solely for proline and a proline/glutamate antiporter, is shown in a functional study in which we found the following: (1) Mitochondria undergo passive swelling in isotonic proline solutions in a stereospecific manner. (2) Externally added L: -proline (Pro) generates a mitochondrial membrane potential (Delta Psi) with a rate depending on the transport of Pro across the mitochondrial inner membrane. (3) The dependence of the rate of generation of Delta Psi on increasing Pro concentrations exhibits hyperbolic kinetics. Proline transport is inhibited in a competitive manner by the non-penetrant thiol reagent mersalyl, but it is insensitive to the penetrant thiol reagent N-ethylmaleimide (NEM). (4) No accumulation of proline occurs inside the mitochondria as a result of the addition of proline externally, whereas the content of glutamate increases both in mitochondria and in the extramitochondrial phase. (5) Glutamate efflux from mitochondria occurs at a rate which depends on the mitochondrial transport, and it is inhibited in a non-competitive manner by NEM. The dependence of the rate of glutamate efflux on increasing proline concentration shows saturation kinetics. The physiological role of carrier-mediated transport in the regulation of proline catabolism, as well as the possible occurrence of a proline/glutamate shuttle in durum wheat seedlings mitochondria, are discussed.

Unraveling Δ1-Pyrroline-5-Carboxylate-Proline Cycle in Plants by Uncoupled Expression of Proline Oxidation Enzymes

“…Therefore, its further reduction to Pro depends on P5CR availability and the possible existence of yet undefined P5C transporter(s) in the mitochondrial outer membrane. Until now only a Pro-specific inward transporter and Pro (inward)/Glu (outward) antiporter have been identified in the mitochondrial membrane of durum wheat, responsible for Pro entrance (66). However, these transporters have not been tested for their ability to transfer P5C.…”

Improving salinity resilience inSwertia chirayitaclonal line withLactobacillus plantarum

“…A modified method described by Costilow and Cooper (1978) was carried out to assay the activity of proline dehydrogenase. The enzyme reaction mixture containing 100 mM sodium carbonate buffer (pH 10.3), 20 mM L-proline solution and 10 mM NAD was used.…”

“…During mitochondrial oxidative phosphorylation, oxidation of proline may provide reductants for energy synthesis (ATP) (Hare and Cress 1997). The presence of specific amino acid transporters like the proline uniporter and the proline/glutamate indicated the active process of proline uptake into the mitochondria (Di Martino et al 2006). High levels of proline synthesis in the cytosol during stress may sustain normal levels of NAD(P) + /NAD(P)H ratio (Hare and Cress 1997).…”

Metabolic and molecular-genetic regulation of proline signaling and itscross-talk with major effectors mediates abiotic stress tolerance in plants

“…The mitochondria of Triticum durum contains a Pro uniporter, which aids in the passage of Pro into the mitochondrial matrix. A Pro-glutamate shuttle between the cytosol and the mitochondrial matrix has also been identified in a Pro/ glutamate antiporter in the mitochondrial membrane of T. durum (Di Martino et al, 2006). The delivery of arginine and ornithine through the mitochondrial membrane is sustained by the basic amino acid (BAC) transporters (Palmieri et al, 2006).…”