Functional Characterization of Betaine/Proline Transporters in Betaine-accumulating Mangrove

Abstract

Betaine is an important osmoprotectant in many plants, but its transport activity has only been demonstrated using a proline transporter from tomato, a betaine-nonaccumulating plant. In this study, two fulllength and one partial transporter genes were isolated from betaine-accumulating mangrove Avicennia marina. Their homologies to betaine transporters from bacteria and betaine/4-aminobutyrate transporters from mammalian cells were low but were high to proline transporters from Arabidopsis and tomato. Two fulllength transporters could complement the Na ؉ -sensitive phenotype of the Escherichia coli mutant deficient in betT, putPA, proP, and proU. Both transporters could efficiently take up betaine and proline with similar affinities (K m , 0.32-0.43 mM) and maximum velocities (1.9 -3.6 nmol/min/mg of protein). The uptakes of betaine and proline were significantly inhibited by mono-and dimethylglycine but only partially inhibited by betaine aldehyde, choline, and 4-aminobutyrate. Sodium and potassium chloride markedly enhanced betaine uptake rates with optimum concentrations at 0.5 M, whereas sucrose showed only modest activation. The change of amino acids Thr 290 -Thr-Ser 292 in a putative periplasmic loop to Arg 290 -Gly-Arg 292 yielded the active transporter independent of salts, suggesting the positive charge induced a conformational change to the active form. These data clearly indicate that the betaine-accumulating mangrove contains betaine/proline transporters whose properties are distinct from betaine transporters of bacteria and mammalian cells.

The AtProT Family. Compatible Solute Transporters with Similar Substrate Specificity But Differential Expression Patterns

“…In general, for all three transporters, higher affinity correlated with lower capacity, and vice versa. For the ProT homologs AmT1 and AmT2 from mangrove, sodium and potassium ions have been shown to enhance transport activity (Waditee et al, 2002). However, no increased AtProT-mediated transport rates could be measured by the addition of sodium chloride to the assay medium (data not shown).…”

“…. However, the affinity of AtProT2 and AtProT1 for Gly betaine, a substrate for LeProT1 from tomato and the ProT homologs from mangrove, as well as the kinetic properties of AtProT3, had not been determined so far Waditee et al, 2002). Transport assays showed that all three AtProTs were able to mediate uptake of labeled Pro, Gly betaine, and GABA.…”

“…The steady-state mRNA levels of the Arabidopsis ProT2 and the three mangrove ProT homologs (AmT1-3) increased under salt stress conditions, as do the concentrations of Pro and Gly betaine, respectively (Waditee et al, 2002). Furthermore, HvProT expression in barley roots was increased by salt stress treatments (Ueda et al, 2001).…”

“…AAF76897), and mangrove (Avicennia marina; AmT1-3; Waditee et al, 2002). Originally, the transporters were described as specific Pro transporters (ProTs; Rentsch et al, 1996).…”

“…Originally, the transporters were described as specific Pro transporters (ProTs; Rentsch et al, 1996). However, ProTs from tomato and the betaine-accumulating mangrove A. marina were later shown to be general transporters for the compatible solutes Pro and Gly betaine Waditee et al, 2002). Furthermore, the Arabidopsis ProT2 was shown to mediate transport of Pro and the stress-induced compound g-aminobutyric acid (GABA), although GABA was recognized with lower affinity .…”