A central role for polyprenol reductase in plant dolichol biosynthesis

•Metabolic engineering of dolichol biosynthesis in Nicotiana benthamiana.•Polyprenol reductase is the bottleneck step in dolichol biosynthesis.•Over-expressing three enzymes, CPT3, CPTBP and PPRD increases dolichol.•Long-chain polyprenols are increased by 400-fold. Dolichol is an essential polyisopr...

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Published in:	Plant science (Limerick) Vol. 303; p. 110773
Main Authors:	Van Gelder, Kristen, Virta, Lilia K.A., Easlick, Jeremy, Prudhomme, Nicholas, McAlister, Jason A., Geddes-McAlister, Jennifer, Akhtar, Tariq A.
Format:	Journal Article
Language:	English
Published:	Ireland Elsevier B.V 01-02-2021
Subjects:	cis-Prenyltransferase cis-Prenyltransferase binding protein Dolichol Dolichols - biosynthesis Endoplasmic reticulum Metabolic Networks and Pathways Nicotiana - enzymology Nicotiana - genetics Nicotiana - metabolism Oxidoreductases - genetics Oxidoreductases - metabolism Phylogeny Plant Proteins - genetics Plant Proteins - metabolism Polyisoprenoid Polyprenol Polyprenol reductase Solanum lycopersicum - enzymology Solanum lycopersicum - genetics Solanum lycopersicum - metabolism Transferases - metabolism GGPP GPP FPP IPP cis-Prenyltransferase Polyisoprenoid cis-Prenyltransferase binding protein Polyprenol reductase Dolichol Polyprenol C5 CPTs SlCPT3 MVA MEP SlCPTBP DMAPP TPTs Endoplasmic reticulum
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Summary:	•Metabolic engineering of dolichol biosynthesis in Nicotiana benthamiana.•Polyprenol reductase is the bottleneck step in dolichol biosynthesis.•Over-expressing three enzymes, CPT3, CPTBP and PPRD increases dolichol.•Long-chain polyprenols are increased by 400-fold. Dolichol is an essential polyisoprenoid within the endoplasmic reticulum of all eukaryotes. It serves as a membrane bound anchor onto which N-glycans are assembled prior to being transferred to nascent polypeptides, many of which enter the secretory pathway. Historically, it has been posited that the accumulation of dolichol represents the ‘rate-limiting’ step in the evolutionary conserved process of N-glycosylation, which ultimately affects the efficacy of approximately one fifth of the entire eukaryotic proteome. Therefore, this study aimed to enhance dolichol accumulation by manipulating the enzymes involved in its biosynthesis using an established Nicotiana benthamiana platform. Co-expression of a Solanum lycopersicum (tomato) cis-prenyltransferase (CPT) and its cognate partner protein, CPT binding protein (CPTBP), that catalyze the antepenultimate step in dolichol biosynthesis led to a 400-fold increase in the levels of long-chain polyprenols but resulted in only modest increases in dolichol accumulation. However, when combined with a newly characterized tomato polyprenol reductase, dolichol biosynthesis was enhanced by approximately 20-fold. We provide further evidence that in the aquatic macrophyte, Lemna gibba, dolichol is derived exclusively from the mevalonic acid (MVA) pathway with little participation from the evolutionary co-adopted non-MVA pathway. Taken together these results indicate that to effectively enhance the in planta accumulation of dolichol, coordinated synthesis and reduction of polyprenol to dolichol, is strictly required.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0168-9452 1873-2259
DOI:	10.1016/j.plantsci.2020.110773