Functional characterization of squalene synthase and squalene epoxidase in Taraxacum koksaghyz

Abstract

The Russian dandelion Taraxacum koksaghyz produces high-value isoprenoids such as pentacyclic triterpenes and natural rubber in the latex of specialized cells known as

Uncovering mechanisms of rubber biosynthesis in Taraxacum koksaghyz – role of cis‐prenyltransferase‐like 1 protein

“…Remarkably, some putative enzymes of the MEP pathway were significantly ( P < 0.01) upregulated in the TkCPTL1 ‐RNAi lines: Comparing the mean values of log2 LFQ intensities of all replicates of the four transgenic lines (#15) to the mean value of wild‐type plants (#4), resulted differences of 1.134 for 4‐hydroxy‐3‐methylbut‐2‐enyl diphosphate reductase (HDR) as well as 0.621 and 0.9785 for 2‐C‐methyl‐ d ‐erythritol 2,4‐cyclodiphosphate synthase (MCS) and 4‐hydroxy‐3‐methylbut‐2‐en‐1‐yl diphosphate synthase (HDS), respectively. Downstream of the branching point farnesyl pyrophosphate (FPP), the enzyme squalene epoxidase (SQE; also named as squalene monooxygenase, see Table ), catalyzing the epoxidation of squalene to 2,3‐oxidosqualene, was increased in latex of TkCPTL1 ‐RNAi lines compared with wild‐type (Unland et al ., ). 2,3‐oxidosqualene functions as precursor to either pentacyclic triterpenes or sterols.…”

“…2,3‐oxidosqualene functions as precursor to either pentacyclic triterpenes or sterols. Other proteins that were less abundant in TkCPTL1 knockdowns included two oxidosqualene cyclases (OSCs) responsible for converting of 2,3‐oxidosqualene to several pentacyclic triterpene products (Unland et al ., ; Pütter et al ., ). Interestingly, REF and SRPPs exhibited very high protein levels in the wild‐type and remained unaffected in the RNAi lines, as it was indicated in the SRRP‐specific western blot analysis (Figure f).…”

“…Additionally, in our proteomic approach, we observed that SQE was upregulated as well (Figure ). Along with SQS, this enzyme is a rate‐limiting enzyme in triterpene synthesis and provides 2,3‐oxidosqualene for cyclization reactions catalyzed by several OSCs such as TkOSC1, a mixed cyclase that produces the major pentacyclic triterpenes in dandelion (Unland et al ., ). These particular pentacyclic triterpenes were substantially increased in the latex fractions of TkCPTL1 ‐RNAi lines.…”

“…Thus, in these lines, SQE was possibly upregulated to channel more molecules toward triterpene generation. Interestingly, two OSCs were downregulated, even though triterpene levels were elevated and a co‐regulation with TkSQE1 was previously described in TkSQE1 ‐RNAi plants (Figure ) (Unland et al ., ). As such, it is possible that cells can sense when a certain triterpene amount is present and then they synthesize these enzymes in a feedback‐regulated way.…”

“…Triterpenes were extracted from 100 mg of lyophilized and ground root material as specified before (Unland et al ., ).…”