Expression of a bacterial 3-dehydroshikimate dehydratase (QsuB) reduces lignin and improves biomass saccharification efficiency in switchgrass (Panicum virgatum L.)

Abstract

Background Lignin deposited in plant cell walls negatively affects biomass conversion into advanced bioproducts. There is therefore a strong interest in developing bioenergy crops with reduced lignin content or altered lignin structures. Another desired trait for bioenergy crops is the ability to accumulate novel bioproducts, which would enhance the development of economically sustainable biorefineries. As previously demonstrated in the model plant Arabidopsis, expression of a 3-dehydroshikimate dehydratase in plants offers the potential for decreasing lignin content and overproducing a value-added metabolic coproduct (i.e., protocatechuate) suitable for biological upgrading. Results The 3-dehydroshikimate dehydratase QsuB from Corynebacterium glutamicum was expressed in the bioenergy crop switchgrass (Panicum virgatum L.) using the stem-specific promoter of an O-methyltransferase gene (pShOMT) from sugarcane. The activity of pShOMT was validated in switchgrass after observation in-situ of beta-glucuronidase (GUS) activity in stem nodes of plants carrying a pShOMT::GUS fusion construct. Under controlled growth conditions, engineered switchgrass lines containing a pShOMT::QsuB construct showed reductions of lignin content, improvements of biomass saccharification efficiency, and accumulated higher amount of protocatechuate compared to control plants. Attempts to generate transgenic switchgrass lines carrying the QsuB gene under the control of the constitutive promoter pZmUbi-1 were unsuccessful, suggesting possible toxicity issues associated with ectopic QsuB expression during the plant regeneration process. Conclusion This study validates the transfer of the QsuB engineering approach from a model plant to switchgrass. We have demonstrated altered expression of two important traits: lignin content and accumulation of a co-product. We found that the choice of promoter to drive QsuB expression should be carefully considered when deploying this strategy to other bioenergy crops. Field-testing of engineered QsuB switchgrass are in progress to assess the performance of the introduced traits and agronomic performances of the transgenic plants.

Expression of Dehydroshikimate Dehydratase in Sorghum Improves Biomass Yield, Accumulation of Protocatechuate, and Biorefinery Economics

“…6,35 Overall, the DHBA titers measured in QsuB sorghum biomass are approximately 10-fold higher than those measured previously in switchgrass modified with the qsuB gene, but lower than those from QsuB poplar in which DHBA ranged between 0.7−1.2% on a DW basis in woody biomass. 18,19 Besides differences in crop types, the use of different promoters to drive qsuB expression in these crops can explain the variability observed for the DHBA content.…”

A new approach to zip‐lignin: 3,4‐dihydroxybenzoate is compatible with lignification

“…Gain-offunction approaches that not only reduce cell wall recalcitrance but which also impart additional value to biomass represent a particularly promising strategy to enable more cost-effective processing of woody biomass. In this study, we transformed hybrid poplar with a bacterial 3dehydroshikimate dehydratase gene (QsuB) that was previously shown to reduce lignin deposition and improve saccharification yields in Arabidopsis and switchgrass (Eudes et al, 2015;Hao et al, 2021). Expression of QsuB in hybrid poplar led to the hyperaccumulation of soluble DHB in the xylem, and effectively diverted carbon flux away from the phenylpropanoid pathway resulting in decreased lignin content, altered lignin monomer composition, and improved saccharification potential compared to WT trees.…”

“…Employing the same lignin‐specific promoter, the QsuB ‐poplar lines in this study produced up to 11.5 mg g −1 of extractable DHBA in xylem tissue (Fig. 1) compared to just 1.45, 0.133 and 0.375 mg g −1 produced by QsuB ‐expressing tobacco, Arabidopsis, and switchgrass, respectively (Eudes et al ., 2015; Wu et al ., 2017; Hao et al ., 2021). Given the fast growth rates of hybrid poplar and the proven capacity to generate substantial amounts of DHB, these trees could prove to be useful as value‐added woody feedstocks.…”

Engineering sorghum for higher 4-hydroxybenzoic acid content

“…Total RNAs were extracted from the 5-cm bottom part of the main stem from plants in the T2 generation at three different developmental stages using the RNeasy Plant Mini Kit (Qiagen, Redwood City, CA). cDNA synthesis was conducted using the SuperScript IV First-Strand Synthesis kit (Thermo Fisher Scientific, Waltham, MA) as previously described ( Hao et al, 2021 ). RT-qPCR was performed using 35 cycles consisting of 5 s at 95 °C for denaturation and 15 s at 60 °C for annealing and amplification.…”