Plant Physiology 97:317-321 (1991)
© 1991 American Society of Plant Biologists
Molecular Biology and Gene Regulation
Cloning and Sequencing of the cDNA Encoding the Rubber Elongation Factor of Hevea brasiliensis
Elisabeth Goyvaerts,
Mark Dennis,
David Light1 and
Nam-Hai Chua
Institute of Molecular and Cell Biology, National University of Singapore, Singapore 0511,
Department of Medicinal and Biomolecular Chemistry, Genentech, Inc., South San Francisco, California 94080,
Laboratory of Plant Molecular Biology, The Rockefeller University, New York, New York 10021-6396
In Hevea brasiliensis, the rubber particle in the laticiferous vessel is the site of rubber (cis-1-4-polyisoprene) biosynthesis. A 14 kilodalton protein, rubber elongation factor (REF), is associated with the rubber particle in a ratio of one REF to one rubber molecule (Dennis M, Henzel W, Bell J, Kohr W, Light D [1989] J Biol Chem 264: 18618-18628; Dennis M, Light D [1989] J Biol Chem 264: 18608-18617). To obtain more information concerning the function of REF and its synthesis and assembly in the rubber particle, we isolated cDNA clones encoding REF. We used antibodies to REF to screen a Hevea leaf  gt11 cDNA expression library and obtained several positive clones. Sequence analysis of the REF cDNA clones showed that the REF mRNA contains 121 nucleotides of 5'-nontranslated sequences and a 205 nucleotide 3'-nontranslated region. The open reading frame encodes the entire 14 kilodalton REF protein without any extra amino acids (Dennis M, Henzel W, Bell J, Kohr W, Light D [1989] J Biol Chem 264: 18618-18628). The REF cDNA was subcloned in pGEM-3Z/-4Z and expressed in vitro. The translation product is a 14 kilodalton protein that can be immunoprecipitated with antibodies to REF. Addition of microsomal membranes to the in vitro translation product did not alter the mobility of the REF protein. This, and the sequence data, indicate that REF is not made as a preprotein. Our results suggest that REF is synthesized on free polysomes in the laticifer cytoplasm and that assembly of the rubber particles is likely to occur in the cytosol.
1 Present address: Codon, 213 East Grand Avenue, South San Francisco, CA 94080.
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