Gene silencing of mannose 6-phosphate reductase in the parasitic weed Orobanche aegyptiaca through the production of homologous dsRNA sequences in the host plant.
Orobanche spp. (broomrape) are parasitic plants which subsist on the roots of a wide range of hosts, including tomato, causing severe losses in yield quality and quantity. Large amounts of mannitol accumulate in this parasitic weed during development. Mannose 6-phosphate reductase (M6PR) is a key enzyme in mannitol biosynthesis, and it has been suggested that mannitol accumulation may be very important for Orobanche development. Therefore, the Orobanche M6PR gene is a potential target for efforts to control this parasite. Transgenic tomato plants were produced bearing a gene construct containing a specific 277-bp fragment from Orobanche aegyptiaca M6PR-mRNA, in an inverted-repeat configuration. M6PR-siRNA was detected in three independent transgenic tomato lines in the R1 generation, but was not detected in the parasite. Quantitative RT-PCR analysis showed that the amount of endogenous M6PR mRNA in the tubercles and underground shoots of O. aegyptiaca grown on transgenic host plants was reduced by 60%-80%. Concomitant with M6PR mRNA suppression, there was a significant decrease in mannitol level and a significant increase in the percentage of dead O. aegyptiaca tubercles on the transgenic host plants. The detection of mir390, which is involved with cytoplasmic dsRNA processing, is the first indication of the existence of gene-silencing mechanisms in Orobanche spp. Gene silencing mechanisms are probably involved with the production of decreased levels of M6PR mRNA in the parasites grown on the transformed tomato lines.
Aly
Radi
R
Department of Plant Pathology and Weed Research, ARO, The Volcani Center, Newe-Yaar Research Center, Ramat Yishay, Israel. radi@volcani.agri.gov.il
Cholakh
Hila
H
Joel
Daniel M
DM
Leibman
Diana
D
Steinitz
Benjamin
B
Zelcer
Aaron
A
Naglis
Anna
A
Yarden
Oded
O
Gal-On
Amit
A
D001483:Base Sequence; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D006790:Host-Parasite Interactions / Q000235:genetics*; D018551:Solanum lycopersicum / Q000235:genetics / Q000469:parasitology*; D008969:Molecular Sequence Data; D031672:Orobanche / Q000201:enzymology / Q000235:genetics*; D018517:Plant Roots / Q000235:genetics / Q000469:parasitology; D030821:Plants, Genetically Modified / Q000235:genetics / Q000469:parasitology; D012330:RNA, Double-Stranded / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D018749:RNA, Plant / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D016415:Sequence Alignment; D013401:Sugar Alcohol Dehydrogenases / Q000235:genetics*
D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D018749:RNA, Plant; D034741:RNA, Small Interfering; D013401:Sugar Alcohol Dehydrogenases; C076442:mannose-6-phosphate reductase