Natural rubber (NR) is an important material widely used in industry. Unfortunately, the global market is predicted to experience a severe NR shortage, primarily due to increases in consumption of NR. Currently, rubber production is dependent upon a single source, the para-rubber tree (Hevea brasiliensis). However, the relatively limited rubber production currently available from rubber trees has highlighted the need to develop alternative sources of rubber. The desert shrub guayule (Parthenium argentatum) provides an attractive source for natural rubber, particularly if latex yields from this plant can be increased via metabolic engineering. Based on preliminary data, we hypothesize that rubber synthesis in guayule and other rubber-producing plants is catalyzed by an enzyme complex (Rubber Synthetase Complex; RSC) located in the membrane of the rubber particle (the cellular organelle in which rubber synthesis occurs). The goal of the proposed research is to functionally characterize putative members of the RSC, with the long-term objective of metabolically engineering rubber production in both plant (guayule), microbial, and synthetic production systems. The objectives of this project are to: (1) identify membrane-associated members of RSC; (2) functionally characterize individual RSC members and reconstituted RSC complexes in microbial or synthetic systems; (3) increase rubber production in guayule through metabolic engineering of RSC. The proposed research will improve the fundamental understanding of how rubber production occurs in plant systems, and will provide a foundation for the generation of both alternative rubber production systems, as well as synthetic systems designed to produce custom polymers.