Date of this Version
Plant Biotechnology Journal
The impact of metabolic engineering on nontarget pathways and outcomes of metabolic engineering from different genomes are poorly understood questions. Therefore, squalene biosynthesis genes FARNESYL DIPHOSPHATE SYNTHASE (FPS) and SQUALENE SYNTHASE (SQS) were engineered via the Nicotiana tabacum chloroplast (C), nuclear (N) or both (CN) genome to promote squalene biosynthesis. SQS levels were ~4300-fold higher in C and CN lines than in N, but all accumulated ~150-fold higher squalene due to substrate or storage limitations. Abnormal leaf and flower phenotypes, including lower pollen production and reduced fertility, were observed regardless of the compartment or level of transgene expression. Substantial changes in metabolomes of all lines were observed: levels of 65-120 unrelated metabolites, including the toxic alkaloid nicotine, changed by as much as 32-fold. Profound effects of transgenesis on nontarget gene expression included changes in the abundance of 19 076 transcripts by up to 2000-fold in CN; 7784 transcripts by up to 1400-fold in N; and 5224 transcripts by as much as 2200-fold in C. Transporter-related transcripts were induced, and cell cycle-associated transcripts were disproportionately repressed in all three lines. Transcriptome changes were validated by qRT-PCR. The mechanism underlying these large changes likely involves matabolite-mediated anterograde and/or retrograde signalling irrespective of the level of transgene expression or end product, due to imbalance of metabolic pools, offering new insight into both anticipated and unanticipated consequences of metabolic engineering.
© Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd All articles accepted from 1st January 2016 (date of flip) are published under the terms of the Creative Commons Attribution License. Articles accepted before this date were published under the agreement as stated in the final article.
secondary metabolism, retrograde/anterograde signalling, transgenic/transplastomic plants, squalene
Pasoreck, E. K., Su, J., Silverman, I. M., Gosai, S. J., Gregory, B. D., Yuan, J. S., & Daniell, H. (2016). Terpene Metabolic Engineering via Nuclear or Chloroplast Genomes Profoundly and Globally Impacts Off-Target Pathways Through Metabolite Signalling. Plant Biotechnology Journal, 14 (9), 1862-1875. http://dx.doi.org/10.1111/pbi.12548
Additional FilesFigS1_Terpene Metabolic Engineering_2016.pdf (69 kB)
FigS2_Terpene Metabolic Engineering_2016.pdf (135 kB)
FigS3_Terpene Metabolic Engineering_2016.pdf (92 kB)
FigS4_Terpene Metabolic Engineering_2016.pdf (115 kB)
TableS1_Terpene Metabolic Engineering_2016.xlsx (25 kB)
TableS2_Terpene Metabolic Engineering_2016.xlsx (13 kB)
TableS3_Terpene Metabolic Engineering_2016.pdf (207 kB)
Date Posted: 14 July 2017
This document has been peer reviewed.