The Para rubber tree (Hevea brasiliensis) is an economically important tropical tree species that produces natural rubber, an essential industrial raw material. Here we present a high-quality genome assembly of this species (1.37 Gb, scaffold N50 = 1.28 Mb) that covers 93.8% of the genome (1.47 Gb) and harbours 43,792 predicted protein-coding genes. A striking expansion of the REF/SRPP (rubber elongation factor/small rubber particle protein) gene family and its divergence into several laticifer-specific isoforms seem crucial for rubber biosynthesis. The REF/SRPP family has isoforms with sizes similar to or larger than SRPP1 (204 amino acids) in 17 other plants examined, but no isoforms with similar sizes to REF1 (138 amino acids), the predominant molecular variant. A pivotal point in Hevea evolution was the emergence of REF1, which is located on the surface of large rubber particles that account for 93% of rubber in the latex (despite constituting only 6% of total rubber particles, large and small). The stringent control of ethylene synthesis under active ethylene signalling and response in laticifers resolves a longstanding mystery of ethylene stimulation in rubber production. Our study, which includes the re-sequencing of five other Hevea cultivars and extensive RNA-seq data, provides a valuable resource for functional genomics and tools for breeding elite Hevea cultivars.

Biochemical evidence reported so far suggests that rubber synthesis takes place on the surface of rubber particles suspended in the latex of Hevea brasiliensis. We have isolated and characterized a cDNA clone that encodes a protein tightly bound on a small rubber particle. We named this protein small rubber particle protein (SRPP). Prior to this study, this protein was known as a latex allergen, and only its partial amino acid sequence was reported. Sequence analysis revealed that this protein is highly homologous to the rubber elongation factor and the Phaseolus vulgaris stress-related protein. Southern and Northern analyses indicate that the protein is encoded by a single gene and highly expressed in latex. An allergenicity test using the recombinant protein confirmed that the cloned cDNA encodes the known 24-kDa latex allergen. Neither ethylene stimulation nor wounding changed the transcript level of the SRPP gene in H. brasiliensis. An in vitro rubber assay showed that the protein plays a positive role in rubber biosynthesis. Therefore, it is likely that SRPP is a part of the rubber biosynthesis machinery, if not the rubber polymerase, along with the rubber elongation factor.Rubber (cis-1,4-polyisoprene), an isoprenoid polymer with no known physiological function to the plant, is produced in about 2000 plant species with varying degrees of quality and quantity (1). Rubber is the raw material of choice for heavy duty tires and other industrial uses requiring elasticity, flexibility, and resilience. Hevea brasiliensis has been the only commercial source of natural rubber mainly because of its abundance in the tree, its quality, and the ease of harvesting. The diminishing acreage of rubber plantations and life-threatening latex allergy to Hevea rubber, coupled with an increasing demand, have prompted research interests in the study of rubber biosynthesis and the development of alternative rubber sources.In H. brasiliensis, rubber synthesis takes place on the surface of rubber particles suspended in the latex (the cytoplasm of laticifers). The laticifers are specialized vessels that are located adjacent to the phloem of the rubber tree. When severed during tapping, the high turgor pressure inside the laticifers expels latex containing 30 -50% (w/w) cis-1,4-polyisoprene. The latex can be fractionated by centrifugation into three phases: the top fraction containing mostly rubber particles, the metabolically active middle fraction (called C-serum), and the bottom fraction of mainly vacuole-like organelles called lutoids. More than 240 expressed sequence tags (ESTs) 1 have been identified from the latex of H. brasiliensis 2 Kush et al. (2) have shown differential expression of several rubber biosynthesis-related genes in latex. The rubber elongation factor (REF), an enzyme involved in rubber biosynthesis (3), is highly expressed in laticifers (4). Laticiferous cells actively translate the transcribed genes into proteins. About 200 distinct polypeptides are present in the latex of H. brasiliensis (5). Arokiaraj et al...

Hevea brasiliensis is the most widely cultivated species for commercial production of natural rubber (cis-polyisoprene). In this study, 10,040 expressed sequence tags (ESTs) were generated from the latex of the rubber tree, which represents the cytoplasmic content of a single cell type, in order to analyse the latex transcription profile with emphasis on rubber biosynthesis-related genes. A total of 3,441 unique transcripts (UTs) were obtained after quality editing and assembly of EST sequences. Functional classification of UTs according to the Gene Ontology convention showed that 73.8% were related to genes of unknown function. Among highly expressed ESTs, a significant proportion encoded proteins related to rubber biosynthesis and stress or defence responses. Sequences encoding rubber particle membrane proteins (RPMPs) belonging to three protein families accounted for 12% of the ESTs. Characterization of these ESTs revealed nine RPMP variants (7.9-27 kDa) including the 14 kDa REF (rubber elongation factor) and 22 kDa SRPP (small rubber particle protein). The expression of multiple RPMP isoforms in latex was shown using antibodies against REF and SRPP. Both EST and quantitative reverse transcription-PCR (QRT-PCR) analyses demonstrated REF and SRPP to be the most abundant transcripts in latex. Besides rubber biosynthesis, comparative sequence analysis showed that the RPMPs are highly similar to sequences in the plant kingdom having stress-related functions. Implications of the RPMP function in cis-polyisoprene biosynthesis in the context of transcript abundance and differential gene expression are discussed.

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