A Lettuce (Lactuca sativa) Homolog of Human Nogo-B Receptor Interacts with cis-Prenyltransferase and Is Necessary for Natural Rubber Biosynthesis

Natural rubber ( cis -1,4-polyisoprene) is an indispensable biopolymer used to manufacture diverse consumer products. Although a major source of natural rubber is the rubber tree ( Hevea brasiliensis ), lettuce ( Lactuca sativa ) is also known to synthesize natural rubber. Here, we report that an unusual cis -prenyltransferase-like 2 ( CPTL2 ) that lacks the conserved motifs of conventional cis -prenyltransferase is required for natural rubber biosynthesis in lettuce. CPTL2 , identified from the lettuce rubber particle proteome, displays homology to a human NogoB receptor and is predominantly expressed in latex. Multiple transgenic lettuces expressing CPTL2-RNAi constructs showed that a decrease of CPTL2 transcripts (3–15% CPTL2 expression relative to controls) coincided with the reduction of natural rubber as low as 5%. We also identified a conventional cis -prenyltransferase 3 ( CPT3 ), exclusively expressed in latex. In subcellular localization studies using fluorescent proteins, cytosolic CPT3 was relocalized to endoplasmic reticulum by co-occurrence of CPTL2 in tobacco and yeast at the log phase. Furthermore, yeast two-hybrid data showed that CPTL2 and CPT3 interact. Yeast microsomes containing CPTL2/CPT3 showed enhanced synthesis of short cis -polyisoprenes, but natural rubber could not be synthesized in vitro . Intriguingly, a homologous pair CPTL1/CPT1, which displays ubiquitous expressions in lettuce, showed a potent dolichol biosynthetic activity in vitro . Taken together, our data suggest that CPTL2 is a scaffolding protein that tethers CPT3 on endoplasmic reticulum and is necessary for natural rubber biosynthesis in planta , but yeast-expressed CPTL2 and CPT3 alone could not synthesize high molecular weight natural rubber in vitro .Natural rubber biosynthesis in lettuce ( Lactuca sativa ) and other plants remains elusive. Results An unusual cis -prenyltransferase-like protein interacts with and tethers a cis -prenyltransferase on endoplasmic reticulum, and its RNAi-silencing eliminates natural rubber. Conclusion cis -Prenyltransferase-like protein is a necessary component in natural rubber biosynthesis in lettuce. Significance The results presented here suggest hetero-protein complexes are involved in natural rubber biosynthesis.