RNAi-mediated replacement of morphine with the nonnarcotic alkaloid reticuline in opium poppy.
We report on the silencing of codeinone reductase (COR) in the opium poppy, Papaver somniferum, using a chimeric hairpin RNA construct designed to silence all members of the multigene COR family through RNA interference (RNAi). After gene silencing, the precursor alkaloid (S)-reticuline-seven enzymatic steps upstream of codeinone-accumulated in transgenic plants at the expense of morphine, codeine, oripavine and thebaine. Methylated derivatives of reticuline also accumulated. Analysis verified loss of Cor gene transcript, appearance of 22-mer degradation products and reduction of enzyme activity. The surprising accumulation of (S)-reticuline suggests a feedback mechanism preventing intermediates from general benzylisoquinoline synthesis entering the morphine-specific branch. However transcript levels for seven other enzymes in the pathway, both before and after (S)-reticuline, were unaffected. This is the first report of gene silencing in transgenic opium poppy and of metabolic engineering to cause the high-yield accumulation of the nonnarcotic alkaloid reticuline.
Allen
Robert S
RS
CSIRO Plant Industry, PO Box 1600, Canberra, Australian Capital Territory 2601, Australia.
Millgate
Anthony G
AG
Chitty
Julie A
JA
Thisleton
Jennifer
J
Miller
James A C
JA
Fist
Anthony J
AJ
Gerlach
Wayne L
WL
Larkin
Philip J
PJ
D000429:Alcohol Oxidoreductases / Q000172:deficiency* / Q000235:genetics; D000470:Alkaloids / Q000235:genetics / Q000378:metabolism*; D044182:Benzylisoquinolines / Q000378:metabolism*; D002478:Cells, Cultured; D005818:Genetic Engineering / Q000379:methods*; D009020:Morphine / Q000378:metabolism*; D064430:NAD (+) and NADP (+) Dependent Alcohol Oxidoreductases; D010207:Papaver / Q000235:genetics* / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference
D000470:Alkaloids; D044182:Benzylisoquinolines; D009020:Morphine; D000429:Alcohol Oxidoreductases; D064430:NAD (+) and NADP (+) Dependent Alcohol Oxidoreductases; C096678:codeinone reductase (NADPH); C003298:reticuline