Evolution of the isoprene biosynthetic pathway in kudzu.

Isoprene synthase converts dimethylallyl diphosphate, derived from the methylerythritol 4-phosphate (MEP) pathway, to isoprene. Isoprene is made by some plants in substantial amounts, which affects atmospheric chemistry, while other plants make no isoprene. As part of our long-term study of isoprene synthesis, the genetics of the isoprene biosynthetic pathway of the isoprene emitter, kudzu (Pueraria montana), was compared with similar genes in Arabidopsis (Arabidopsis thaliana), which does not make isoprene. The MEP pathway genes in kudzu were similar to the corresponding Arabidopsis genes. Isoprene synthase genes of kudzu and aspen (Populus tremuloides) were cloned to compare their divergence with the divergence seen in MEP pathway genes. Phylogenetic analysis of the terpene synthase gene family indicated that isoprene synthases are either within the monoterpene synthase clade or sister to it. In Arabidopsis, the gene most similar to isoprene synthase is a myrcene/ocimene (acyclic monoterpenes) synthase. Two phenylalanine residues found exclusively in isoprene synthases make the active site smaller than other terpene synthase enzymes, possibly conferring specificity for the five-carbon substrate rather than precursors of the larger isoprenoids. Expression of the kudzu isoprene synthase gene in Arabidopsis caused Arabidopsis to emit isoprene, indicating that whether or not a plant emits isoprene depends on whether or not it has a terpene synthase capable of using dimethylallyl diphosphate.

Genes encoding chitinase-antifreeze proteins are regulated by cold and expressed by all cell types in winter rye shoots.

One group of antifreeze proteins (AFPs) is composed of two chitinases that accumulate in the apoplast of winter rye leaves during cold acclimation. In this study, the 28- and 35-kDa chitinase-AFPs were localized in nonacclimated and cold-acclimated rye leaves by immunoelectron microscopy with an antiserum produced against the purified winter rye 35-kDa chitinase-AFP. In cold-acclimated winter rye leaves, labelled chitinase-AFPs were abundant in the walls of epidermal, parenchymal sheath and mesophyll cells and xylem vessels, while less label was present in walls of vascular parenchyma cells. In contrast, chitinase labelling was essentially absent in the nonacclimated cells except in xylem vessels. As shown by RNA blotting, the transcripts of chitinase-AFPs accumulated to a high level in rye leaves during cold acclimation, to a lesser extent in crowns and were not detectable in roots. mRNA transcripts of the 28-kDa chitinase-AFP were localized in rye leaves by in situ hybridization. The chitinase-AFP transcripts were found in the same cell types as the protein itself. We conclude that all metabolically active cell types in cold-acclimated winter rye leaves and crowns are able to synthesize chitinase-AFPs and secrete them into adjacent cell walls, where they may interact with ice to delay its propagation through the plant and modify its growth.

ISOPRENE EMISSION FROM PLANTS.

Very large amounts of isoprene are emitted from vegetation, especially from mosses, ferns, and trees. This hydrocarbon flux to the atmosphere, roughly equal to the flux of methane, has a large effect on the oxidizing potential of the atmosphere. Isoprene emission results from de novo synthesis by the deoxyxylulose phosphate/methyl erythritol 4-phosphate pathway in plastids. Dimethylallyl pyrophosphate made by this pathway is converted to isoprene by isoprene synthase. Isoprene synthase activity in plants has a high pH optimum and requirement for Mg2+ that is consistent with its location inside chloroplasts. Isoprene emission costs the plant significant amounts of carbon, ATP, and reducing power. Researchers hypothesize that plants benefit from isoprene emission because it helps photosynthesis recover from short high-temperature episodes. The evolution of isoprene emission may have been important in allowing plants to survive the rapid temperature changes that can occur in air because of the very low heat capacity of isoprene relative to water.