Abstract
BackgroundPlant cell suspension cultures can be used for the production of valuable pharmaceutical and industrial proteins. When the recombinant protein is secreted into the culture medium, restricting expression to a defined growth phase can improve both the quality and quantity of the recovered product by minimizing proteolytic activity. Temporal restriction is also useful for recombinant proteins whose constitutive expression affects cell growth and viability, such as viral interleukin-10 (vIL-10).ResultsWe have developed a novel, tetracycline-inducible system suitable for tobacco BY-2 suspension cells which increases the yields of vIL-10. The new system is based on a binary vector that is easier to handle than conventional vectors, contains an enhanced inducible promoter and 5′-UTR to improve yields, and incorporates a constitutively-expressed visible marker gene to allow the rapid and straightforward selection of the most promising transformed clones. Stable transformation of BY-2 cells with this vector, without extensive optimization of the induction conditions, led to a 3.5 fold increase in vIL-10 levels compared to constitutive expression in the same host.ConclusionsWe have developed an effective and straightforward molecular farming platform technology that improves both the quality and the quantity of recombinant proteins produced in plant cells, particularly those whose constitutive expression has a negative impact on plant growth and development. Although we tested the platform using vIL-10 produced in BY-2 cells, it can be applied to other host/product combinations and is also useful for basic research requiring strictly controlled transgene expression.
Highlights
Plant cell suspension cultures can be used for the production of valuable pharmaceutical and industrial proteins
We recently reported the expression of the viral antiinflammatory cytokine interleukin-10 in transgenic tobacco plants, and found that retention of the recombinant protein in the endoplasmic reticulum (ER) resulted in stunted plant growth, with the severity of the phenotype determined by the level of protein accumulation [14 and Additional file 1: Figure S1]
There was no significant difference in the growth properties of the cell suspension cultures based on pellet weight, but it was clear that viral interleukin-10 (vIL-10) was produced at much lower levels than mIL-10
Summary
Plant cell suspension cultures can be used for the production of valuable pharmaceutical and industrial proteins. Molecular farming is the production of valuable pharmaceutical or industrial proteins in whole plants and plantbased systems such as cell suspension cultures [1,2,3,4,5,6]. Inducible promoters are often used to restrict transgene expression to particular developmental stages in whole plants, or to particular growth phases in cultivated cells, because this can improve product quality and quantity. We recently reported the expression of the viral antiinflammatory cytokine interleukin-10 (vIL-10) in transgenic tobacco plants, and found that retention of the recombinant protein in the endoplasmic reticulum (ER) resulted in stunted plant growth, with the severity of the phenotype determined by the level of protein accumulation [14 and Additional file 1: Figure S1]. We decided to express vIL-10 in the tobacco cell line Bright Yellow 2 (BY-2) and to develop strategies based on temporally-restricted inducible expression to avoid the deleterious effects observed in planta
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