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Metabolic Engineering of Squalene Biosynthesis via Different Genomes Impacts Non-target Cellular Metabolomic and Transcriptomic Pathways

ABSTRACT: The impact of metabolic engineering on non-target pathways is a poorly explored question that requires deeper mechanistic investigation. Therefore, Nicotiana tabacum was engineered via the chloroplast (C), nuclear (N) or both (CN) genomes to express genes encoding FARNESYL DIPHOSPHATE SYNTHASE (FPS) and SQUALENE SYNTHASE (SQS) to promote squalene biosynthesis. SQS levels were ~4,300-fold higher in C and CN lines than in the N line, but all accumulated ~150-fold more squalene due to substrate or storage limitations. Slower growth and abnormal flowering phenotypes occurred regardless of the compartment or level of transgene expression. Substantial changes in metabolomes of all lines were observed: 65-120 unrelated metabolites changed > 32-fold. Profound effects of transgenesis on non-target gene expression included changes in the expression of 19,076 transcripts by > 2,000-fold in the CN line; 7,784 transcripts by > 1,400-fold in the N line; and 5,224 transcripts by > 2,200-fold in the C line. Cell cycle-associated transcripts were disproportionally repressed in all three lines, as shown by both manual and Gene Ontology analysis, and transporter-related transcripts were induced. Trends observed in transcriptomes were further validated by qRT-PCR. The mechanism underlying these large changes likely involves metabolite-mediated anterograde and/or retrograde signaling.

ORGANISM(S): Nicotiana tabacum

PROVIDER: GSE74103 | GEO | 2016/06/05

SECONDARY ACCESSION(S): PRJNA299013

REPOSITORIES: GEO

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