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Sucrose metabolism gene families and their biological functions.

Jiang SY, Chi YH, Wang JZ, Zhou JX, Cheng YS, Zhang BL, Ma A, Vanitha J, Ramachandran S - Sci Rep (2015)

Bottom Line: Although studies on general metabolism pathway were well documented, less information is available on the genome-wide identification of these genes, their expansion and evolutionary history as well as their biological functions.They were evolutionarily conserved under purifying selection among species and expression divergence played important roles for gene survival after expansion.Overexpression of 15 sorghum genes in Arabidopsis revealed their roles in biomass accumulation, flowering time control, seed germination and response to high salinity and sugar stresses.

View Article: PubMed Central - PubMed

Affiliation: Genome Structural Biology Group, Temasek Life Sciences Laboratory, National University of Singapore, Singapore 117604.

ABSTRACT
Sucrose, as the main product of photosynthesis, plays crucial roles in plant development. Although studies on general metabolism pathway were well documented, less information is available on the genome-wide identification of these genes, their expansion and evolutionary history as well as their biological functions. We focused on four sucrose metabolism related gene families including sucrose synthase, sucrose phosphate synthase, sucrose phosphate phosphatase and UDP-glucose pyrophosphorylase. These gene families exhibited different expansion and evolutionary history as their host genomes experienced differentiated rates of the whole genome duplication, tandem and segmental duplication, or mobile element mediated gene gain and loss. They were evolutionarily conserved under purifying selection among species and expression divergence played important roles for gene survival after expansion. However, we have detected recent positive selection during intra-species divergence. Overexpression of 15 sorghum genes in Arabidopsis revealed their roles in biomass accumulation, flowering time control, seed germination and response to high salinity and sugar stresses. Our studies uncovered the molecular mechanisms of gene expansion and evolution and also provided new insight into the role of positive selection in intra-species divergence. Overexpression data revealed novel biological functions of these genes in flowering time control and seed germination under normal and stress conditions.

No MeSH data available.


Related in: MedlinePlus

Molecular and phenotypic characterization of transgenic Arabidopsis plants overexpressing sweet sorghum SPS genes.(A) Expression patterns of 4 SPS genes in sweet sorghum Keller by RT-PCR analysis. Total RNA samples were prepared from 7 different tissues. YR, young root, from 14-day seedlings; MR, mature root, from 3-month plants; S, stem, from 30-day plants; YL, young leaf, from 14-day seedlings; ML, mature leaf, from 3-month plants; YP, young panicle, from un-flowering panicles; MP, mature panicle, from flowering panicles. The prefix “Sobic.” was omitted in each locus name. (B) Three of five sorghum SPS genes were independently overexpressed in Arabidopsis under the maize ubiquitin promoter to generate at least 4 independent transgenic lines with single copy of T-DNA insertion in each construct. NA, not available. (C) An example of copy number detection by Southern blot hybridization. DNA samples from 11 transgenic plant (ubiquitin:: Sobic.009G233200) were digested by PstI for the hybridization using the probe prepared from the hygromycin gene. (D) Expression analysis of 3 independent transgenic plants (ubiquitin::Sobic.009G233200) with single copy of T-DNA insertion. ACT1, an Arabidopsis gene encoding Actin-1 protein with locus name At2g37620. (E–G) Effects of sucrose, glucose and maltose treatments, respectively, on germination rates of WT and transgenic plants. D2, D3, D4, D5, D6, D8 and D10 indicated 2, 3, 4, 5, 6, 8 and 10 days after inoculation on ½ MS media. (H–J) Phenotypic observation of transgenic plants treated by sucrose, glucose and maltose, respectively, by comparing with WT.
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f7: Molecular and phenotypic characterization of transgenic Arabidopsis plants overexpressing sweet sorghum SPS genes.(A) Expression patterns of 4 SPS genes in sweet sorghum Keller by RT-PCR analysis. Total RNA samples were prepared from 7 different tissues. YR, young root, from 14-day seedlings; MR, mature root, from 3-month plants; S, stem, from 30-day plants; YL, young leaf, from 14-day seedlings; ML, mature leaf, from 3-month plants; YP, young panicle, from un-flowering panicles; MP, mature panicle, from flowering panicles. The prefix “Sobic.” was omitted in each locus name. (B) Three of five sorghum SPS genes were independently overexpressed in Arabidopsis under the maize ubiquitin promoter to generate at least 4 independent transgenic lines with single copy of T-DNA insertion in each construct. NA, not available. (C) An example of copy number detection by Southern blot hybridization. DNA samples from 11 transgenic plant (ubiquitin:: Sobic.009G233200) were digested by PstI for the hybridization using the probe prepared from the hygromycin gene. (D) Expression analysis of 3 independent transgenic plants (ubiquitin::Sobic.009G233200) with single copy of T-DNA insertion. ACT1, an Arabidopsis gene encoding Actin-1 protein with locus name At2g37620. (E–G) Effects of sucrose, glucose and maltose treatments, respectively, on germination rates of WT and transgenic plants. D2, D3, D4, D5, D6, D8 and D10 indicated 2, 3, 4, 5, 6, 8 and 10 days after inoculation on ½ MS media. (H–J) Phenotypic observation of transgenic plants treated by sucrose, glucose and maltose, respectively, by comparing with WT.

Mentions: In sorghum, we have identified a total of 5 SPS genes. In most of cases, these genes were expressed in multiple tissues (Fig. 7A). However, in each gene, the expression abundance varied among different tissues. Three genes including Sobic.003G403300, Sobic.004G068400 and Sobic.010G205100 showed the highest transcript level at young leaves and the gene Sobic.009G233200 exhibited the highest expression abundance in stem (Fig. 7A). To better understand their biological functions, coding regions from three genes were amplified from sweet sorghum by RT-PCR for overexpression under the maize ubiquitin promoter in Arabidopsis (Fig. 7B). By molecular characterization, we identified 13, 4, and 15 independent T2 lines from the genes Sobic.004G068400, Sobic.009G233200 and Sobic.010G205100, respectively, which showed high expression level with single copy of T-DNA insertion (Fig. 7B). These lines were then subjected into phenotypic characterization. For both genes Sobic.004G068400 and Sobic.010G205100, no obvious phenotypic variation was observed after overexpression in Arabidopsis when compared with WT plants. Here we focused on the gene Sobic.009G233200. Based on Southern blot hybridization, we have selected 4 independent lines with Single copy of T-DNA insertion (Fig. 7C). These lines showed high expression level for the overexpressed gene in Arabidopsis (Fig. 7D). We then subjected these lines into morphological characterization. Our data showed that overexpression of Sobic.009G233200 significantly retarded seed germination (Fig. 7E–G and Supplementary Fig. S3 A-C). Under normal growth conditions, up to 93% of Arabidopsis WT seeds germinated after 2-day inoculation on ½ MS media whereas significantly lower percentages of seeds germinated for all identified 4 independent transgenic lines. Here we showed the results from two lines indicated by red and green cylinders, respectively in the Fig. 7E–G; Supplementary Figure S3 A-C. Their germination rates in these two lines were only 66% and 35%, respectively, under normal growth media (Supplementary Fig. S3A). Under 1–9% of sucrose stress, germination for both WT and transgenic seeds were significantly retarded. However, transgenic seeds showed more sensitive response to the stress. For example, after 7-day inoculation on 7% sucrose containing MS media, up to 80% of WT seeds germinated whereas only less than 10% seeds germinated for the transgenic lines (Fig. 7E,H). Similarly, germination retardation was also observed for the transgenic seeds when they were germinated under both glucose and maltose stresses (Fig. 7 F and G; Supplementary Fig. S3 B and C). Although the transgenic Arabidopsis plants showed retarded germination under either normal or sugar stress conditions during early days, these lines exhibited no significant difference after more days of incubation on ½ MS media (Fig. 7I,J). From two weeks after germination, we further measured plant height, root length, flowering time and biomass yield during the whole developmental stages and found that no statistical difference was detected between WT and transgenic lines.


Sucrose metabolism gene families and their biological functions.

Jiang SY, Chi YH, Wang JZ, Zhou JX, Cheng YS, Zhang BL, Ma A, Vanitha J, Ramachandran S - Sci Rep (2015)

Molecular and phenotypic characterization of transgenic Arabidopsis plants overexpressing sweet sorghum SPS genes.(A) Expression patterns of 4 SPS genes in sweet sorghum Keller by RT-PCR analysis. Total RNA samples were prepared from 7 different tissues. YR, young root, from 14-day seedlings; MR, mature root, from 3-month plants; S, stem, from 30-day plants; YL, young leaf, from 14-day seedlings; ML, mature leaf, from 3-month plants; YP, young panicle, from un-flowering panicles; MP, mature panicle, from flowering panicles. The prefix “Sobic.” was omitted in each locus name. (B) Three of five sorghum SPS genes were independently overexpressed in Arabidopsis under the maize ubiquitin promoter to generate at least 4 independent transgenic lines with single copy of T-DNA insertion in each construct. NA, not available. (C) An example of copy number detection by Southern blot hybridization. DNA samples from 11 transgenic plant (ubiquitin:: Sobic.009G233200) were digested by PstI for the hybridization using the probe prepared from the hygromycin gene. (D) Expression analysis of 3 independent transgenic plants (ubiquitin::Sobic.009G233200) with single copy of T-DNA insertion. ACT1, an Arabidopsis gene encoding Actin-1 protein with locus name At2g37620. (E–G) Effects of sucrose, glucose and maltose treatments, respectively, on germination rates of WT and transgenic plants. D2, D3, D4, D5, D6, D8 and D10 indicated 2, 3, 4, 5, 6, 8 and 10 days after inoculation on ½ MS media. (H–J) Phenotypic observation of transgenic plants treated by sucrose, glucose and maltose, respectively, by comparing with WT.
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Related In: Results  -  Collection

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f7: Molecular and phenotypic characterization of transgenic Arabidopsis plants overexpressing sweet sorghum SPS genes.(A) Expression patterns of 4 SPS genes in sweet sorghum Keller by RT-PCR analysis. Total RNA samples were prepared from 7 different tissues. YR, young root, from 14-day seedlings; MR, mature root, from 3-month plants; S, stem, from 30-day plants; YL, young leaf, from 14-day seedlings; ML, mature leaf, from 3-month plants; YP, young panicle, from un-flowering panicles; MP, mature panicle, from flowering panicles. The prefix “Sobic.” was omitted in each locus name. (B) Three of five sorghum SPS genes were independently overexpressed in Arabidopsis under the maize ubiquitin promoter to generate at least 4 independent transgenic lines with single copy of T-DNA insertion in each construct. NA, not available. (C) An example of copy number detection by Southern blot hybridization. DNA samples from 11 transgenic plant (ubiquitin:: Sobic.009G233200) were digested by PstI for the hybridization using the probe prepared from the hygromycin gene. (D) Expression analysis of 3 independent transgenic plants (ubiquitin::Sobic.009G233200) with single copy of T-DNA insertion. ACT1, an Arabidopsis gene encoding Actin-1 protein with locus name At2g37620. (E–G) Effects of sucrose, glucose and maltose treatments, respectively, on germination rates of WT and transgenic plants. D2, D3, D4, D5, D6, D8 and D10 indicated 2, 3, 4, 5, 6, 8 and 10 days after inoculation on ½ MS media. (H–J) Phenotypic observation of transgenic plants treated by sucrose, glucose and maltose, respectively, by comparing with WT.
Mentions: In sorghum, we have identified a total of 5 SPS genes. In most of cases, these genes were expressed in multiple tissues (Fig. 7A). However, in each gene, the expression abundance varied among different tissues. Three genes including Sobic.003G403300, Sobic.004G068400 and Sobic.010G205100 showed the highest transcript level at young leaves and the gene Sobic.009G233200 exhibited the highest expression abundance in stem (Fig. 7A). To better understand their biological functions, coding regions from three genes were amplified from sweet sorghum by RT-PCR for overexpression under the maize ubiquitin promoter in Arabidopsis (Fig. 7B). By molecular characterization, we identified 13, 4, and 15 independent T2 lines from the genes Sobic.004G068400, Sobic.009G233200 and Sobic.010G205100, respectively, which showed high expression level with single copy of T-DNA insertion (Fig. 7B). These lines were then subjected into phenotypic characterization. For both genes Sobic.004G068400 and Sobic.010G205100, no obvious phenotypic variation was observed after overexpression in Arabidopsis when compared with WT plants. Here we focused on the gene Sobic.009G233200. Based on Southern blot hybridization, we have selected 4 independent lines with Single copy of T-DNA insertion (Fig. 7C). These lines showed high expression level for the overexpressed gene in Arabidopsis (Fig. 7D). We then subjected these lines into morphological characterization. Our data showed that overexpression of Sobic.009G233200 significantly retarded seed germination (Fig. 7E–G and Supplementary Fig. S3 A-C). Under normal growth conditions, up to 93% of Arabidopsis WT seeds germinated after 2-day inoculation on ½ MS media whereas significantly lower percentages of seeds germinated for all identified 4 independent transgenic lines. Here we showed the results from two lines indicated by red and green cylinders, respectively in the Fig. 7E–G; Supplementary Figure S3 A-C. Their germination rates in these two lines were only 66% and 35%, respectively, under normal growth media (Supplementary Fig. S3A). Under 1–9% of sucrose stress, germination for both WT and transgenic seeds were significantly retarded. However, transgenic seeds showed more sensitive response to the stress. For example, after 7-day inoculation on 7% sucrose containing MS media, up to 80% of WT seeds germinated whereas only less than 10% seeds germinated for the transgenic lines (Fig. 7E,H). Similarly, germination retardation was also observed for the transgenic seeds when they were germinated under both glucose and maltose stresses (Fig. 7 F and G; Supplementary Fig. S3 B and C). Although the transgenic Arabidopsis plants showed retarded germination under either normal or sugar stress conditions during early days, these lines exhibited no significant difference after more days of incubation on ½ MS media (Fig. 7I,J). From two weeks after germination, we further measured plant height, root length, flowering time and biomass yield during the whole developmental stages and found that no statistical difference was detected between WT and transgenic lines.

Bottom Line: Although studies on general metabolism pathway were well documented, less information is available on the genome-wide identification of these genes, their expansion and evolutionary history as well as their biological functions.They were evolutionarily conserved under purifying selection among species and expression divergence played important roles for gene survival after expansion.Overexpression of 15 sorghum genes in Arabidopsis revealed their roles in biomass accumulation, flowering time control, seed germination and response to high salinity and sugar stresses.

View Article: PubMed Central - PubMed

Affiliation: Genome Structural Biology Group, Temasek Life Sciences Laboratory, National University of Singapore, Singapore 117604.

ABSTRACT
Sucrose, as the main product of photosynthesis, plays crucial roles in plant development. Although studies on general metabolism pathway were well documented, less information is available on the genome-wide identification of these genes, their expansion and evolutionary history as well as their biological functions. We focused on four sucrose metabolism related gene families including sucrose synthase, sucrose phosphate synthase, sucrose phosphate phosphatase and UDP-glucose pyrophosphorylase. These gene families exhibited different expansion and evolutionary history as their host genomes experienced differentiated rates of the whole genome duplication, tandem and segmental duplication, or mobile element mediated gene gain and loss. They were evolutionarily conserved under purifying selection among species and expression divergence played important roles for gene survival after expansion. However, we have detected recent positive selection during intra-species divergence. Overexpression of 15 sorghum genes in Arabidopsis revealed their roles in biomass accumulation, flowering time control, seed germination and response to high salinity and sugar stresses. Our studies uncovered the molecular mechanisms of gene expansion and evolution and also provided new insight into the role of positive selection in intra-species divergence. Overexpression data revealed novel biological functions of these genes in flowering time control and seed germination under normal and stress conditions.

No MeSH data available.


Related in: MedlinePlus