Biochemical characterization of rice trehalose‐6‐phosphate phosphatases supports distinctive functions of these plant enzymes

Abstract

Substantial levels of trehalose accumulate in bacteria, fungi, and invertebrates, where it serves as a storage carbohydrate or as a protectant against environmental stresses. In higher plants, trehalose is detected at fairly low levels; therefore, a regulatory or signaling function has been proposed for this molecule. In many organisms, trehalose‐6‐phosphate phosphatase is the enzyme governing the final step of trehalose biosynthesis. Here we report that OsTPP1 and OsTPP2 are the two major trehalose‐6‐phosphate phosphatase genes expressed in vegetative tissues of rice. Similar to results obtained from our previous OsTPP1 study, complementation analysis of a yeast trehalose‐6‐phosphate phosphatase mutant and activity measurement of the recombinant protein demonstrated that OsTPP2 encodes a functional trehalose‐6‐phosphate phosphatase enzyme. OsTPP2 expression is transiently induced in response to chilling and other abiotic stresses. Enzymatic characterization of recombinant OsTPP1 and OsTPP2 revealed stringent substrate specificity for trehalose 6‐phosphate and about 10 times lower Km values for trehalose 6‐phosphate as compared with trehalose‐6‐phosphate phosphatase enzymes from microorganisms. OsTPP1 and OsTPP2 also clearly contrasted with microbial enzymes, in that they are generally unstable, almost completely losing activity when subjected to heat treatment at 50 °C for 4 min. These characteristics of rice trehalose‐6‐phosphate phosphatase enzymes are consistent with very low cellular substrate concentration and tightly regulated gene expression. These data also support a plant‐specific function of trehalose biosynthesis in response to environmental stresses.

Trehalose and Abiotic Stress in Biological Systems

“…Rice OsTPP1 and OsTPP2 are transiently induced by cold, salt and drought stress as well as external ABA applications (Pramanik & Imai, 2005;Shima et al, 2007). Trehalose was also transiently induced following chilling stress, and its accumulation coincided with the phase change of glucose and fructose levels (Pramanik and Imai 2005).…”

“…Rice OsTPP1 and OsTPP2 are transiently induced by cold, salt and drought stress as well as external ABA applications (Pramanik & Imai, 2005;Shima et al, 2007). Garcia et al (1997) reported as well trehalose accumulation in small amounts in rice roots 3 days following salt stress.…”

Trehalose Metabolites in Arabidopsis—elusive, active and central

“…Only one gene encoding trehalase is present, AtTRE1 (Muller et al, 2001). Therefore radiation at the level of T6P rather than trehalose has occurred in Arabidopsis, and this holds true for other plants with sequenced genomes (Avonce et al, 2006;Lunn, 2007;Pramanik and Imai, 2005;Satoh-Nagasawa et al, 2006;Shima et al, 2007).…”

“…The activities were specific for T6P and did not cleave sucrose-6-phosphate or G6P. In rice, OsTPP1 and OsTPP2 have been characterized biochemically: recombinant enzymes showed a high specificity for T6P and worked at higher affinity with 10 fold lower K m compared to microbial enzymes (Pramanik and Imai, 2005;Shima et al, 2007). Maize RA3 (encoded by RAMOSA3) is a TPP shown to complement S. cerevisiae defective in ScTPS2 and its recombinant protein also cleaves T6P specifically (Satoh-Nagasawa et al, 2006).…”

“…Trehalose is cleaved into two moieties of glucose by specific trehalases (TreH pathway). Plants have been shown to encode enzymes capable of trehalose synthesis via the TPS/TPP pathway (Blazquez et al, 1998;Chary et al, 2007;Pramanik and Imai, 2005;Satoh-Nagasawa et al, 2006;Shima et al, 2007;Vogel et al, 1998). Plants also contain genes that encode trehalase (Aeschbacher et al, 1999;Muller et al, 2001), but lack genes with homology to enzymes from the other pathways.…”