Horizontal Transfer of a Synthetic Metabolic Pathway between Plant Species

Abstract

Transgene expression from the plastid (chloroplast) genome provides unique advantages, including high levels of foreign protein accumulation, convenient transgene stacking in operons, and increased biosafety due to exclusion of plastids from pollen transmission [1, 2]. However, applications in biotechnology and synthetic biology are severely restricted by the very small number of plant species whose plastid genomes currently can be transformed [3, 4]. Here we report a simple method for the introduction of useful plastid transgenes into non-transformable species. The transgenes tested comprised a synthetic operon encoding three components of a biosynthetic pathway for producing the high-value ketocarotenoid astaxanthin in the plastids of the cigarette tobacco, Nicotiana tabacum. Transplastomic N. tabacum plants accumulated astaxanthin to up to 1% of the plants' dry weight. We then used grafting, a procedure recently shown to facilitate horizontal genome transfer between plants [5-7], to let the transgenic chloroplast genome move across the graft junction from N. tabacum plants into plants of the nicotine-free tree species Nicotiana glauca. Transplastomic N. glauca trees expressing the synthetic pathway were recovered at high frequency, thus providing a straightforward method for extension of the transplastomic technology to new species.

The road to astaxanthin production in tomato fruit reveals plastid and metabolic adaptation resulting in an unintended high lycopene genotype with delayed over‐ripening properties

“…In the homozygous, state loss of vigour leading to unviable plants occurred, presumably as a consequence of high non‐endogenous carotenoid levels interfering with plant processes. Reduced vigour on ketocarotenoid production has been reported previously following transplastomic approaches and subsequent horizontal genome transfer to new species (Lu et al ., ). Industrially, a robust phenotype in a single or double hemizygous state (e.g.…”

Non‐endogenous ketocarotenoid accumulation in engineeredSynechocystissp. PCC 6803

“…Carotenoids play a crucial role in photosynthetic organisms and any significant change in their abundance or composition can in principle exert detrimental effects on cell growth and fitness, as reported, for example, for Asx‐accumulating transplastomic tobacco plants which showed a reduced growth rate (Hasunuma et al , Lu et al ). For this reason, in the present study, a temperature‐inducible expression system has been used for the metabolic engineering of the carotenoid biosynthetic pathway in Synechocystis (Fig.…”

A global perspective on carotenoids: Metabolism, biotechnology, and benefits for nutrition and health

“…Since the demonstration that plant leaves could be used to produce non-native ketocarotenoid pigments such as astaxanthin [240][241][242], many efforts have been focused on diverting the endogenous carotenoid pathway of several plants towards the biosynthesis of ketocarotenoids in their leaves. Perhaps the most outstanding examples include the generation of transplastomic lines and nuclear transformants of N. tabacum and the nicotine-free tobacco tree species N. glauca [243][244][245]. Transplastomic tobacco (N. tabacum) plants harboring codonoptimized genes from the marine bacterium Brevundimonas encoding β-carotene ketolase (crtW) and β-carotene hydroxylase (crtZ) produced up to 5 mg/g DW of astaxanthin, reaching more than 0.5% dry weight [243].…”