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The carotenogenesis pathway via the isoprenoid-beta-carotene interference approach in a new strain of Dunaliella salina isolated from Baja California Mexico.

Paniagua-Michel J, Capa-Robles W, Olmos-Soto J, Gutierrez-Millan LE - Mar Drugs (2009)

Bottom Line: When Carotenogenic and non-carotenogenic cells of D. salina BC02 were grown under photoautotrophic growth conditions in the presence of 200 microM fosmidomycin, carotenogenesis and the synthesis of beta-carotene were interrupted after two days in cultured D. salina cells.This result is an indirect consequence of the inhibition of the synthesis of isoprenoids and activity of the recombinant DXR enzyme thereby preventing the conversion of 1-deoxy-D-xylulose 5-phosphate (DXP) to 2-C-methyl-D-erythritol (MEP) and consequently interrupts the early steps of carotenogenesis in D. salina.The effect at the level of proteins and RNA was not evident.

View Article: PubMed Central - PubMed

Affiliation: Department of Marine Biotechnology, Centro de Investigación Científica y de Educación Superior de Ensenada, Ensenada 22860, México. jpaniagu@cicese.mx

ABSTRACT
D. salina is one of the recognized natural sources to produce beta-carotene, and an useful model for studying the role of inhibitors and enhancers of carotenogenesis. However there is little information in D. salina regarding whether the isoprenoid substrate can be influenced by stress factors (carotenogenic) or selective inhibitors which in turn may further contribute to elucidate the early steps of carotenogenesis and biosynthesis of beta-carotene. In this study, Dunaliella salina (BC02) isolated from La Salina BC Mexico, was subjected to the method of isoprenoids-beta-carotene interference in order to promote the interruption or accumulation of the programmed biosynthesis of carotenoids. When Carotenogenic and non-carotenogenic cells of D. salina BC02 were grown under photoautotrophic growth conditions in the presence of 200 microM fosmidomycin, carotenogenesis and the synthesis of beta-carotene were interrupted after two days in cultured D. salina cells. This result is an indirect consequence of the inhibition of the synthesis of isoprenoids and activity of the recombinant DXR enzyme thereby preventing the conversion of 1-deoxy-D-xylulose 5-phosphate (DXP) to 2-C-methyl-D-erythritol (MEP) and consequently interrupts the early steps of carotenogenesis in D. salina. The effect at the level of proteins and RNA was not evident. Mevinolin treated D. salina cells exhibited carotenogenesis and beta-carotene levels very similar to those of control cell cultures indicating that mevinolin not pursued any indirect action in the biosynthesis of isoprenoids and had no effect at the level of the HMG-CoA reductase, the key enzyme of the Ac/MVA pathway.

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HPLC chromatogram of Dunaliella salina cultured in the control growth media (A) after 7 days of culture and in presence of 200 μM fosmidomycin (B) and 1 µM mevinolin (C). The peaks correspond to the following pigments: 1, violaxanthin; 2, neoxanthin; 3, lutein; 4, zeaxanthin; 5, Chlorophyll a; 6, all-trans β-carotene.
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f1-marinedrugs-07-00045: HPLC chromatogram of Dunaliella salina cultured in the control growth media (A) after 7 days of culture and in presence of 200 μM fosmidomycin (B) and 1 µM mevinolin (C). The peaks correspond to the following pigments: 1, violaxanthin; 2, neoxanthin; 3, lutein; 4, zeaxanthin; 5, Chlorophyll a; 6, all-trans β-carotene.

Mentions: Preliminary investigations were carried-out in order to determine the optimal concentration of each inhibitor. Application of fosmidomycin, at a range of concentrations from 50 to 200 μM was assessed in cultured D. salina BCO2 cells. At concentrations below 150 µM, fosmidomycin had no influence on the biosynthesis of β-carotene or on their number of D. salina cells (Table 1), in contrast, when D. salina was exposed to 200 μM fosmidomycin caused growth inhibition and arrested the carotenoids accumulation that was apparent at day 7 of cultivation (Figure 1C). Mevinolin, did not exerted an inhibitory effect on carotenoids accumulation and growth (<5% of control) at a concentration of 1µM, and at 48 h of cultivation as shown in Table 1 and Figure 1B. Once selected the appropriate concentration of each inhibitor, we proceeded to assess its effect on the synthesis of carotenoids, chlorophyll, β-carotene, protein and RNA in D. salina grown under carotenogenic and non-carotenogenic conditions. When D. salina cells were grown under conditions which enhances carotenoids accumulation and in presence of the highest concentration of fosmidomycin (200 μM, the highest response among the assayed concentrations), a decrease in the relative content in carotenoids was registered after four days of culture (Figure 2). This result represented, less than 75% of the content in the cells cultured under non-carotenogenic conditions (control). On the contrary, when D. salina cells were cultured under carotenogenic conditions and exposed to 1 μM mevinolin, this inhibitor did not exerted a significant change in the relative content of carotenoids during the exposure time (four days) when compared to the cells cultured under non-carotenogenic conditions (control) (Figure 2).


The carotenogenesis pathway via the isoprenoid-beta-carotene interference approach in a new strain of Dunaliella salina isolated from Baja California Mexico.

Paniagua-Michel J, Capa-Robles W, Olmos-Soto J, Gutierrez-Millan LE - Mar Drugs (2009)

HPLC chromatogram of Dunaliella salina cultured in the control growth media (A) after 7 days of culture and in presence of 200 μM fosmidomycin (B) and 1 µM mevinolin (C). The peaks correspond to the following pigments: 1, violaxanthin; 2, neoxanthin; 3, lutein; 4, zeaxanthin; 5, Chlorophyll a; 6, all-trans β-carotene.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2666888&req=5

f1-marinedrugs-07-00045: HPLC chromatogram of Dunaliella salina cultured in the control growth media (A) after 7 days of culture and in presence of 200 μM fosmidomycin (B) and 1 µM mevinolin (C). The peaks correspond to the following pigments: 1, violaxanthin; 2, neoxanthin; 3, lutein; 4, zeaxanthin; 5, Chlorophyll a; 6, all-trans β-carotene.
Mentions: Preliminary investigations were carried-out in order to determine the optimal concentration of each inhibitor. Application of fosmidomycin, at a range of concentrations from 50 to 200 μM was assessed in cultured D. salina BCO2 cells. At concentrations below 150 µM, fosmidomycin had no influence on the biosynthesis of β-carotene or on their number of D. salina cells (Table 1), in contrast, when D. salina was exposed to 200 μM fosmidomycin caused growth inhibition and arrested the carotenoids accumulation that was apparent at day 7 of cultivation (Figure 1C). Mevinolin, did not exerted an inhibitory effect on carotenoids accumulation and growth (<5% of control) at a concentration of 1µM, and at 48 h of cultivation as shown in Table 1 and Figure 1B. Once selected the appropriate concentration of each inhibitor, we proceeded to assess its effect on the synthesis of carotenoids, chlorophyll, β-carotene, protein and RNA in D. salina grown under carotenogenic and non-carotenogenic conditions. When D. salina cells were grown under conditions which enhances carotenoids accumulation and in presence of the highest concentration of fosmidomycin (200 μM, the highest response among the assayed concentrations), a decrease in the relative content in carotenoids was registered after four days of culture (Figure 2). This result represented, less than 75% of the content in the cells cultured under non-carotenogenic conditions (control). On the contrary, when D. salina cells were cultured under carotenogenic conditions and exposed to 1 μM mevinolin, this inhibitor did not exerted a significant change in the relative content of carotenoids during the exposure time (four days) when compared to the cells cultured under non-carotenogenic conditions (control) (Figure 2).

Bottom Line: When Carotenogenic and non-carotenogenic cells of D. salina BC02 were grown under photoautotrophic growth conditions in the presence of 200 microM fosmidomycin, carotenogenesis and the synthesis of beta-carotene were interrupted after two days in cultured D. salina cells.This result is an indirect consequence of the inhibition of the synthesis of isoprenoids and activity of the recombinant DXR enzyme thereby preventing the conversion of 1-deoxy-D-xylulose 5-phosphate (DXP) to 2-C-methyl-D-erythritol (MEP) and consequently interrupts the early steps of carotenogenesis in D. salina.The effect at the level of proteins and RNA was not evident.

View Article: PubMed Central - PubMed

Affiliation: Department of Marine Biotechnology, Centro de Investigación Científica y de Educación Superior de Ensenada, Ensenada 22860, México. jpaniagu@cicese.mx

ABSTRACT
D. salina is one of the recognized natural sources to produce beta-carotene, and an useful model for studying the role of inhibitors and enhancers of carotenogenesis. However there is little information in D. salina regarding whether the isoprenoid substrate can be influenced by stress factors (carotenogenic) or selective inhibitors which in turn may further contribute to elucidate the early steps of carotenogenesis and biosynthesis of beta-carotene. In this study, Dunaliella salina (BC02) isolated from La Salina BC Mexico, was subjected to the method of isoprenoids-beta-carotene interference in order to promote the interruption or accumulation of the programmed biosynthesis of carotenoids. When Carotenogenic and non-carotenogenic cells of D. salina BC02 were grown under photoautotrophic growth conditions in the presence of 200 microM fosmidomycin, carotenogenesis and the synthesis of beta-carotene were interrupted after two days in cultured D. salina cells. This result is an indirect consequence of the inhibition of the synthesis of isoprenoids and activity of the recombinant DXR enzyme thereby preventing the conversion of 1-deoxy-D-xylulose 5-phosphate (DXP) to 2-C-methyl-D-erythritol (MEP) and consequently interrupts the early steps of carotenogenesis in D. salina. The effect at the level of proteins and RNA was not evident. Mevinolin treated D. salina cells exhibited carotenogenesis and beta-carotene levels very similar to those of control cell cultures indicating that mevinolin not pursued any indirect action in the biosynthesis of isoprenoids and had no effect at the level of the HMG-CoA reductase, the key enzyme of the Ac/MVA pathway.

Show MeSH