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Higher plant cytochrome b5 polypeptides modulate fatty acid desaturation.

Kumar R, Tran LS, Neelakandan AK, Nguyen HT - PLoS ONE (2012)

Bottom Line: With regard to ω-3 desaturation, certain Cb5 genes of both Arabidopsis and soybean were shown to facilitate the accumulation of more desaturation products than others when co-expressed with their native FAD3.Additionally, similar trends of differential desaturation product accumulation were also observed with most Cb5 genes of both soybean and Arabidopsis even if co-expressed with non-native FAD3.The present study reports the first description of the differential nature of the Cb5 genes of higher plants in fatty acid desaturation and further suggests that ω-3/ω-6 desaturation product outcome is determined by the nature of both the Cb5 isoform and the fatty acid desaturases.

View Article: PubMed Central - PubMed

Affiliation: Division of Plant Sciences and National Center for Soybean Biotechnology, University of Missouri, Columbia, Missouri, United States of America. kraj.pgr@gmail.com

ABSTRACT

Background: Synthesis of polyunsaturated fatty acids (PUFAs) in the endoplasmic reticulum of plants typically involves the fatty acid desaturases FAD2 and FAD3, which use cytochrome b(5) (Cb5) as an electron donor. Higher plants are reported to have multiple isoforms of Cb5, in contrast to a single Cb5 in mammals and yeast. Despite the wealth of information available on the roles of FAD2 and FAD3 in PUFA synthesis, information regarding the contributions of various Cb5 isoforms in desaturase-mediated reactions is limited.

Results: The present functional characterization of Cb5 polypeptides revealed that all Arabidopsis Cb5 isoforms are not similarly efficient in ω-6 desaturation, as evidenced by significant variation in their product outcomes in yeast-based functional assays. On the other hand, characterization of Cb5 polypeptides of soybean (Glycine max) suggested that similar ω-6 desaturation efficiencies were shared by various isoforms. With regard to ω-3 desaturation, certain Cb5 genes of both Arabidopsis and soybean were shown to facilitate the accumulation of more desaturation products than others when co-expressed with their native FAD3. Additionally, similar trends of differential desaturation product accumulation were also observed with most Cb5 genes of both soybean and Arabidopsis even if co-expressed with non-native FAD3.

Conclusions: The present study reports the first description of the differential nature of the Cb5 genes of higher plants in fatty acid desaturation and further suggests that ω-3/ω-6 desaturation product outcome is determined by the nature of both the Cb5 isoform and the fatty acid desaturases.

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The 18∶3 content of yeast co-expressing Cb5 and FAD3 of soybean.A known amount of 18∶2 was added to cultures before induction with galactose and allowed to grow 28°C for 48 h (A) or 15°C for 96 h (B). FAMES were analyzed by GC-FID. Values are expressed as molar percentage of total FAs and represent average and SD of three independent cultures. W = wild type yeast (empty vector pESC control); M = mutant cb5 yeast. See Table S2 for detailed fatty acid composition.
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pone-0031370-g005: The 18∶3 content of yeast co-expressing Cb5 and FAD3 of soybean.A known amount of 18∶2 was added to cultures before induction with galactose and allowed to grow 28°C for 48 h (A) or 15°C for 96 h (B). FAMES were analyzed by GC-FID. Values are expressed as molar percentage of total FAs and represent average and SD of three independent cultures. W = wild type yeast (empty vector pESC control); M = mutant cb5 yeast. See Table S2 for detailed fatty acid composition.

Mentions: In the majority of angiosperms, FAD3 is mainly responsible for ω-3 FA production, primarily 18∶3. Accordingly, the contributions of various Cb5 isoforms in ω-3 desaturation were explored using the soybean, FAD3-1A[39], which is similar to soybean, FAD3B[40]. As expected, in wild type or mutant cb5 yeast transformed with FAD3 and not supplemented with exogenous 18∶2, we failed to detect 18∶3 (Table S2). However, under conditions of 18∶2 feeding, considerable production of 18∶3 was observed in both wild type and mutant cb5 yeast (Figure 5A). Notably, co-expression of soybean native Cb5 and FAD3 genes produced a 2-fold more 18∶3 FAs than expressing FAD3 alone (Figure 5A). While all Cb5 isoforms complemented the defect of cb5 yeast mutant, there was interesting variation in 18∶3 FA accumulation. Although 18∶3 levels were relatively similar in Cb5-C2 and Cb5-E1, both showed ∼1.5-fold more 18∶3 accumulation than Cb5-C3. In Cb5-A1 the 18∶3 level was intermediate to that of Cb5-C2, Cb5-E1 and Cb5-C. At reduced temperature, the levels of ω-3 desaturation product were higher all around, but there was no distinct variation in 18∶3 levels between the Cb5 isoforms (Figure 5B).


Higher plant cytochrome b5 polypeptides modulate fatty acid desaturation.

Kumar R, Tran LS, Neelakandan AK, Nguyen HT - PLoS ONE (2012)

The 18∶3 content of yeast co-expressing Cb5 and FAD3 of soybean.A known amount of 18∶2 was added to cultures before induction with galactose and allowed to grow 28°C for 48 h (A) or 15°C for 96 h (B). FAMES were analyzed by GC-FID. Values are expressed as molar percentage of total FAs and represent average and SD of three independent cultures. W = wild type yeast (empty vector pESC control); M = mutant cb5 yeast. See Table S2 for detailed fatty acid composition.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3285619&req=5

pone-0031370-g005: The 18∶3 content of yeast co-expressing Cb5 and FAD3 of soybean.A known amount of 18∶2 was added to cultures before induction with galactose and allowed to grow 28°C for 48 h (A) or 15°C for 96 h (B). FAMES were analyzed by GC-FID. Values are expressed as molar percentage of total FAs and represent average and SD of three independent cultures. W = wild type yeast (empty vector pESC control); M = mutant cb5 yeast. See Table S2 for detailed fatty acid composition.
Mentions: In the majority of angiosperms, FAD3 is mainly responsible for ω-3 FA production, primarily 18∶3. Accordingly, the contributions of various Cb5 isoforms in ω-3 desaturation were explored using the soybean, FAD3-1A[39], which is similar to soybean, FAD3B[40]. As expected, in wild type or mutant cb5 yeast transformed with FAD3 and not supplemented with exogenous 18∶2, we failed to detect 18∶3 (Table S2). However, under conditions of 18∶2 feeding, considerable production of 18∶3 was observed in both wild type and mutant cb5 yeast (Figure 5A). Notably, co-expression of soybean native Cb5 and FAD3 genes produced a 2-fold more 18∶3 FAs than expressing FAD3 alone (Figure 5A). While all Cb5 isoforms complemented the defect of cb5 yeast mutant, there was interesting variation in 18∶3 FA accumulation. Although 18∶3 levels were relatively similar in Cb5-C2 and Cb5-E1, both showed ∼1.5-fold more 18∶3 accumulation than Cb5-C3. In Cb5-A1 the 18∶3 level was intermediate to that of Cb5-C2, Cb5-E1 and Cb5-C. At reduced temperature, the levels of ω-3 desaturation product were higher all around, but there was no distinct variation in 18∶3 levels between the Cb5 isoforms (Figure 5B).

Bottom Line: With regard to ω-3 desaturation, certain Cb5 genes of both Arabidopsis and soybean were shown to facilitate the accumulation of more desaturation products than others when co-expressed with their native FAD3.Additionally, similar trends of differential desaturation product accumulation were also observed with most Cb5 genes of both soybean and Arabidopsis even if co-expressed with non-native FAD3.The present study reports the first description of the differential nature of the Cb5 genes of higher plants in fatty acid desaturation and further suggests that ω-3/ω-6 desaturation product outcome is determined by the nature of both the Cb5 isoform and the fatty acid desaturases.

View Article: PubMed Central - PubMed

Affiliation: Division of Plant Sciences and National Center for Soybean Biotechnology, University of Missouri, Columbia, Missouri, United States of America. kraj.pgr@gmail.com

ABSTRACT

Background: Synthesis of polyunsaturated fatty acids (PUFAs) in the endoplasmic reticulum of plants typically involves the fatty acid desaturases FAD2 and FAD3, which use cytochrome b(5) (Cb5) as an electron donor. Higher plants are reported to have multiple isoforms of Cb5, in contrast to a single Cb5 in mammals and yeast. Despite the wealth of information available on the roles of FAD2 and FAD3 in PUFA synthesis, information regarding the contributions of various Cb5 isoforms in desaturase-mediated reactions is limited.

Results: The present functional characterization of Cb5 polypeptides revealed that all Arabidopsis Cb5 isoforms are not similarly efficient in ω-6 desaturation, as evidenced by significant variation in their product outcomes in yeast-based functional assays. On the other hand, characterization of Cb5 polypeptides of soybean (Glycine max) suggested that similar ω-6 desaturation efficiencies were shared by various isoforms. With regard to ω-3 desaturation, certain Cb5 genes of both Arabidopsis and soybean were shown to facilitate the accumulation of more desaturation products than others when co-expressed with their native FAD3. Additionally, similar trends of differential desaturation product accumulation were also observed with most Cb5 genes of both soybean and Arabidopsis even if co-expressed with non-native FAD3.

Conclusions: The present study reports the first description of the differential nature of the Cb5 genes of higher plants in fatty acid desaturation and further suggests that ω-3/ω-6 desaturation product outcome is determined by the nature of both the Cb5 isoform and the fatty acid desaturases.

Show MeSH
Related in: MedlinePlus