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Co-overexpression of geraniol-10-hydroxylase and strictosidine synthase improves anti-cancer drug camptothecin accumulation in Ophiorrhiza pumila.

Cui L, Ni X, Ji Q, Teng X, Yang Y, Wu C, Zekria D, Zhang D, Kai G - Sci Rep (2015)

Bottom Line: Overexpression of individual G10H (G lines) significantly improved CPT production with respect to non-transgenic hairy root cultures (NC line) and single STR overexpressing lines (S lines), indicating that G10H plays a more important role in stimulating CPT accumulation than STR in O. pumila.Furthermore, co-overexpression of G10H and STR genes (SG Lines) caused a 56% increase on the yields of CPT compared to NC line and single gene transgenic lines, showed that simultaneous introduction of G10H and STR can produce a synergistic effect on CPT biosynthesis in O. pumila.The MTT assay results indicated that CPT extracted from different lines showed similar anti-tumor activity, suggesting that transgenic O. pumila hairy root lines could be an alternative approach to obtain CPT.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Plant Biotechnology, Development Center of Plant Germplasm Resources, College of Life and Environment Sciences, Shanghai Normal University, Shanghai 200234, China.

ABSTRACT
Camptothecin (CPT) belongs to a group of monoterpenoidindole alkaloids (TIAs) and its derivatives such as irinothecan and topothecan have been widely used worldwide for the treatment of cancer, giving rise to rapidly increasing market demands. Genes from Catharanthus roseus encoding strictosidine synthase (STR) and geraniol 10-hydroxylase (G10H), were separately and simultaneously introduced into Ophiorrhiza pumila hairy roots. Overexpression of individual G10H (G lines) significantly improved CPT production with respect to non-transgenic hairy root cultures (NC line) and single STR overexpressing lines (S lines), indicating that G10H plays a more important role in stimulating CPT accumulation than STR in O. pumila. Furthermore, co-overexpression of G10H and STR genes (SG Lines) caused a 56% increase on the yields of CPT compared to NC line and single gene transgenic lines, showed that simultaneous introduction of G10H and STR can produce a synergistic effect on CPT biosynthesis in O. pumila. The MTT assay results indicated that CPT extracted from different lines showed similar anti-tumor activity, suggesting that transgenic O. pumila hairy root lines could be an alternative approach to obtain CPT. To our knowledge, this is the first report on the enhancement of CPT production in O. pumila employing a metabolic engineering strategy.

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The inhibitory rate of different transgenic line on K562 cells.
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f6: The inhibitory rate of different transgenic line on K562 cells.

Mentions: The anti-tumor activity of CPT extracted from different hairy root lines was estimated by the MTT test in this study. The crude CPT extracts from some selected transgenic O. pumila hairy root lines and wild type O. pumila plant root were used for the MTT test, to compare their anti-tumor activities on the human chronic myelogenous leukemia K562 cell line. The MTT test showed that crude CPT extracts from line S14, G1, SG26 and WT at the same concentration with 6 levels showed similar anti-tumor activity (Fig. 6). When CPT from various sources was normalized at concentrations of 100 ug/ml, the inhibiting rate varied from 64.59% to 68.57%. The MTT assay results showed that there are no significant differences of anti-tumor activity among these transgenic lines and WT lines, and it can be deduced that genetic engineering did not change CPT activity.


Co-overexpression of geraniol-10-hydroxylase and strictosidine synthase improves anti-cancer drug camptothecin accumulation in Ophiorrhiza pumila.

Cui L, Ni X, Ji Q, Teng X, Yang Y, Wu C, Zekria D, Zhang D, Kai G - Sci Rep (2015)

The inhibitory rate of different transgenic line on K562 cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4316170&req=5

f6: The inhibitory rate of different transgenic line on K562 cells.
Mentions: The anti-tumor activity of CPT extracted from different hairy root lines was estimated by the MTT test in this study. The crude CPT extracts from some selected transgenic O. pumila hairy root lines and wild type O. pumila plant root were used for the MTT test, to compare their anti-tumor activities on the human chronic myelogenous leukemia K562 cell line. The MTT test showed that crude CPT extracts from line S14, G1, SG26 and WT at the same concentration with 6 levels showed similar anti-tumor activity (Fig. 6). When CPT from various sources was normalized at concentrations of 100 ug/ml, the inhibiting rate varied from 64.59% to 68.57%. The MTT assay results showed that there are no significant differences of anti-tumor activity among these transgenic lines and WT lines, and it can be deduced that genetic engineering did not change CPT activity.

Bottom Line: Overexpression of individual G10H (G lines) significantly improved CPT production with respect to non-transgenic hairy root cultures (NC line) and single STR overexpressing lines (S lines), indicating that G10H plays a more important role in stimulating CPT accumulation than STR in O. pumila.Furthermore, co-overexpression of G10H and STR genes (SG Lines) caused a 56% increase on the yields of CPT compared to NC line and single gene transgenic lines, showed that simultaneous introduction of G10H and STR can produce a synergistic effect on CPT biosynthesis in O. pumila.The MTT assay results indicated that CPT extracted from different lines showed similar anti-tumor activity, suggesting that transgenic O. pumila hairy root lines could be an alternative approach to obtain CPT.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Plant Biotechnology, Development Center of Plant Germplasm Resources, College of Life and Environment Sciences, Shanghai Normal University, Shanghai 200234, China.

ABSTRACT
Camptothecin (CPT) belongs to a group of monoterpenoidindole alkaloids (TIAs) and its derivatives such as irinothecan and topothecan have been widely used worldwide for the treatment of cancer, giving rise to rapidly increasing market demands. Genes from Catharanthus roseus encoding strictosidine synthase (STR) and geraniol 10-hydroxylase (G10H), were separately and simultaneously introduced into Ophiorrhiza pumila hairy roots. Overexpression of individual G10H (G lines) significantly improved CPT production with respect to non-transgenic hairy root cultures (NC line) and single STR overexpressing lines (S lines), indicating that G10H plays a more important role in stimulating CPT accumulation than STR in O. pumila. Furthermore, co-overexpression of G10H and STR genes (SG Lines) caused a 56% increase on the yields of CPT compared to NC line and single gene transgenic lines, showed that simultaneous introduction of G10H and STR can produce a synergistic effect on CPT biosynthesis in O. pumila. The MTT assay results indicated that CPT extracted from different lines showed similar anti-tumor activity, suggesting that transgenic O. pumila hairy root lines could be an alternative approach to obtain CPT. To our knowledge, this is the first report on the enhancement of CPT production in O. pumila employing a metabolic engineering strategy.

Show MeSH
Related in: MedlinePlus