MOLECULAR ANALYSIS OF DEVELOPMENT TO IMPROVE COTTON FIBER
Location: Cotton Fiber Bioscience Research Unit
Title: Characterization of the mannan synthase promoter from guar (Cyamopsis tetragonoloba)
Submitted to: Plant Cell and Environment
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: January 5, 2011
Publication Date: January 20, 2011
Citation: Naoumkina, M.A., Dixon, R. 2011. Characterization of the mannan synthase promoter from guar (Cyamopsis tetragonoloba). Plant Cell and Environment. (2011)30:997-1006.
Interpretive Summary: Guar (Cyamopsis tetragonaloba L. Taub), a drought-resistant annual legume, is one of the most important commercial sources for seed gums. Guar seed gum, which contains up to 75-85% of galactomannan, is the most cost effective natural thickener. The viscosity and thickening properties of guar gum have found broad applications in the food, cosmetics, paper, pharmaceutical and petroleum industries. The main consumer of guar gum is the petroleum industry, which uses it as an additive to reduce fluid friction during the drilling of oil wells. Guar galactomannan has been preferred to other galactomannan preparations for industrial applications due to its low cost. The desire to increase the viscosity of the gum has led to development of chemical derivatives which, however, are significantly more expensive. The properties of the polymer can potentially be enhanced by genetic modification.
The mechanism of galactomannan biosynthesis has been well studied in the endospermic legumes including fenugreek (Trigonella foenum-graecum L.) and guar (Edwards, et al. 1989; Edwards, et al. 2004; Edwards, et al. 1999; Edwards, et al. 2002; Edwards, et al. 1992; Reid and Bewley 1979; Reid, et al. 2003; Reid, et al. 1992; Reid and Meier 1970; Reid and Meier 1973), where it is synthesized in the seed’s endosperm by the co-action of two membrane-bound enzymes, mannan synthase (MS) and galactosyl transferase. The peak of activity for these two enzymes has been observed exclusively in the endosperm in the later stage of seed maturation around 25 - 35 days after flowering (Dhugga, et al. 2004; Naoumkina, et al. 2007).
The promoter plays the most important role in determining the temporal and spatial expression pattern of a gene, although the final amount of gene product is determined at both transcriptional and post-transcriptional levels. To date, some strong constitutive promoters, such as the cauliflower mosaic virus 35S promoter and the maize ubiquitin promoter are widely used in plant biotechnology research (Battraw and Hall 1990; Christensen, et al. 1992; Saidi, et al. 2009). However, the expression level of the target gene in the desired tissue is often not satisfactory (Drakakaki, et al. 2000). In addition, continuous high expression of a foreign gene in all tissues, or in just non-target tissues, may cause harmful effects in the host plant (Cheon, et al. 2004). For example, use of the bean phaseolin promoter, which is expressed in both embryo and endosperm, to drive a guar MS transgene resulted in increased galactomannan polymer chain length, but this was accompanied by number of off-target effects on sugar metabolism (Naoumkina et al., 2008). Using a strong endosperm-specific promoter to restrict gene expression to only the endosperm may solve such problems.
A number of endosperm-specific promoters, to date only from cereals, have been isolated (Furtado, et al. 2009; Furtado, et al. 2008; Qu le, et al. 2008; Washida, et al. 1999). However, a promoter from a monocotyledonous species can show expression patterns that differ from those of the source organism when tested in eudicotyledonous species, and even in other monocotyledonous species (Furtado, et al. 2008). Possible chromosomal position effects or lack of suitable trans-acting elements in heterologous species have been put forward as reasons for reduced endosperm-specific expression of promoters. The development of endogenous tissue-specific promoters for plant transformation would be one way to avoid this problem.
Isolation of an endogenous endosperm specific promoter will be desirable for genetic engineering of galactomannan metabolism in guar to prevent off-target genetic and metabolic perturbations in the plant. The promoter of the guar MS gene is suitable for this purpose since this gene is highly expressed exclusively in the endosperm tissue of guar seeds. An approximately 1.6 kb gu
Guar seed gum, consisting primarily of a high molecular weight galactomannan, is the most cost effective natural thickener, having broad applications in the food, cosmetics, paper, pharmaceutical and petroleum industries. The properties of the polymer can potentially be enhanced by genetic modification. Development of suitable endosperm specific promoters for use in guar is desirable for metabolic engineering of the seed gum. A ~1.6 kb guar mannan synthase (MS) promoter region has been isolated. The MS promoter sequence was fused with the GUS reporter gene and overexpressed in the heterologous species alfalfa (Medicago sativa). The potential strength and specificity of the MS promoter was compared with those of the constitutive 35S promoter and the seed specific '-phaseolin promoter. Quantitative GUS assays revealed that the MS promoter directs GUS expression specifically in endosperm in transgenic alfalfa. Thus, the guar MS promoter could prove generally useful for directing endosperm-specific expression of transgenes in legume species.