Altered fatty acid composition of Nicotiana benthamiana and Nicotiana excelsior leaves under transient overexpression of the cyanobacterial desC gene

Abstract

Transient heterologous gene expression in two model plant species, Nicotiana benthamiana and N. excelsior, has been used to study the localization of the heterologous Δ9 acyl-lipid desaturase (Δ9 desaturase) of Synechococcus vulcanus in different cell compartments and its functional activity in the cases of the cytosol, chloroplast, and endoplasmic reticulum (ER) localization. The functional activity and substrate specificity of the heterologous desaturase under the conditions of transient expression have been confirmed by comparison of fatty acid (FA) profiles. The Δ9 desaturase, responsible for the synthesis of oleic and palmitoleic acids, has also been shown to strongly promote the accumulation of polyunsaturated FAs. The results convincingly demonstrate that the Δ9 desaturase of the thermophilic cyanobacterium transiently expressed in two Nicotiana species considerably alters lipid metabolism in their leaves towards a higher FA unsaturation. The functional activity of Δ9 desaturase depends on both the model plant species, N. benthamiana or N. excelsior, and the cellular localization of the enzyme. The method of transient expression of heterologous genes in plants is highly effective, inexpensive, and not time-consuming, which makes it attractive for estimating the functional activity and/or substrate specificity of heterologous desaturases.

Transient Gene Expression is an Effective Experimental Tool for the Research into the Fine Mechanisms of Plant Gene Function: Advantages, Limitations, and Solutions

“…The transient expression of Δ9 desaturase in the N. benthamiana and N. excelsior leaves caused a significant decrease in the content of palmitic acid (16:0) as well as an increase in the shares of oleic (18:1), linolenic (18:2), α-linolenic (18:3), and palmitolinolenic (16:3) fatty acids. Moreover, the utility of N. benthamiana and N. excelsior as model plants was convincingly demonstrated in the studies of functional activity and/or substrate specificity of various heterologous desaturases, not only Δ9 desaturase, but also the desaturases converting fatty acids 18:1 into polyunsaturated fatty acids, since the model plants ( N. benthamiana and N. excelsior ) tested in this study have significantly different fatty acid compositions and the C18/C16 ratio [ 55 ].…”

“…It should be emphasized that the appropriate correct controls are extremely important when studying the functions of the genes involved in the plant response to plant pathogens and the defense signaling pathways in plants since the agroinfiltration itself can induce defense responses of the host, interfering with the interpretation of results [ 3 , 4 ]. The plants inoculated with the agrobacteria carrying an empty vector may be an adequate control [ 55 ].…”

Modulation of the Translation Efficiency of Heterologous mRNA and Target Protein Stability in a Plant System: The Case Study of Interferon-αA

“…The transformed bacteria were selected on a medium containing kanamycin and named according to the expression vector used for transformation. The transformed agrobacteria were used for agroinfiltration into the abaxial epidermis of the leaves of 6-week-old N. benthamiana with a syringe without a needle as previously described [ 37 ]. After agroinfiltration, the plants were cultivated under the same conditions for 7 days.…”

A high throughput assay of lichenase activity with Congo red dye in plants

“…The transformed bacteria were selected on a medium containing kanamycin and designated according to the expression vector used for the transformation. The transformed agrobacteria were used for agroinfiltration into the abaxial epidermis of the leaves of 6-week-old N. benthamiana with a syringe without a needle as earlier described [ 21 , 22 ]. After the agroinfiltration, the plants were cultivated under the same conditions for 7 days.…”