Comparative Analysis of the Regulation of Expression and Structures of Two Evolutionarily Divergent Genes for Δ1-Pyrroline-5-Carboxylate Synthetase from Tomato

Abstract

We isolated two tomato (Lycopersicon esculentum) cDNA clones, tomPRO1 and tomPRO2, specifying ⌬ 1 -pyrroline-5-carboxylate synthetase (P5CS), the first enzyme of proline (Pro) biosynthesis. tomPRO1 is unusual because it resembles prokaryotic polycistronic operons (M.G. García-Ríos, T. Fujita, P.C. LaRosa, R.D. Locy, J.M. Clithero, R.A. Bressan, L.N. Csonka [1997] Proc Natl Acad Sci USA 94: 8249-8254), whereas tomPRO2 encodes a fulllength P5CS. We analyzed the accumulation of Pro and the tomPRO1 and tomPRO2 messages in response to NaCl stress and developmental signals. Treatment with 200 mM NaCl resulted in a >60-fold increase in Pro levels in roots and leaves. However, there was a <3-fold increase in the accumulation of the tomPRO2 message and no detectable induction in the level of the tomPRO1 message in response to NaCl stress. Although pollen contained approximately 100-fold higher levels of Pro than other plant tissues, there was no detectable increase in the level of either message in pollen. We conclude that transcriptional regulation of these genes for P5CS is probably not important for the osmotic or pollenspecific regulation of Pro synthesis in tomato. Using restriction fragment-length polymorphism mapping, we determined the locations of tomPRO1 and tomPRO2 loci in the tomato nuclear genome. Sequence comparison suggested that tomPRO1 is similar to prokaryotic P5CS loci, whereas tomPRO2 is closely related to other eukaryotic P5CS genes.

Desensitization of Feedback Inhibition of the Saccharomyces cerevisiae γ-Glutamyl Kinase Enhances Proline Accumulation and Freezing Tolerance

“…This mutation in microbial GKs is novel in that different mutations in other amino acid residues required for the feedback inhibition of GK by proline were identified in bacterial genes that have been studied (11,13,23,30). A similar mutation in the tomato GK (D162N) corresponding to position 154 resulted in the removal of feedback inhibition but a drastic reduction in the specific activity (16). This is probably because the regional sequence similarity in the two GKs is very low.…”

“…Based on the prediction of the secondary structure of S. cerevisiae GK, the sequence is situated in the region between the ␤-sheet (I136 to N140) and the ␣-helix (D156 to I166), which includes a ␤-sheet-like sequence (L145 to K151), of S. cerevisiae GK. Residues 142 to 154, where most of the mutations are concentrated, are mapped to the region that was identified as important for allosteric control of the tomato GK (16). Although the sequence similarity in the regions of the two GKs is low, the regions are suggested to constitute part of the proline-binding site.…”

Proline accumulation and transcriptional regulation of proline biothesynthesis and degradation in Brassica napus

“…In most plant species studied, proline accumulation during stress is the result of reciprocal action of increased biosynthesis and inhibited degradation (6). However, in tomato, dramatic elevation of free proline content during salt stress is not correlated with the expression of either of P5CS genes (7). In Medicago sativa, proline accumulation under salt stress results from the decreased expression of two PDHs rather than the increased expression of P5CS (8).…”

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

“…However, transport of Pro might also be essential under normal growth conditions. Pro accumulation in developing grapevine berries and maturing pollen occurs independently of changes of the key enzyme(s) of Pro biosynthesis, arguing for a different level of regulation or for import of Pro (Fujita et al, 1998;Stines et al, 1999). Additional indications for Pro transport throughout the plant come from transgenic Arabidopsis (Arabidopsis thaliana) lines with reduced Pro biosynthesis (Nanjo et al, 1999), as the morphological alterations in these lines could be rescued by exogenous Pro application.…”