Switchgrass (Panicum virgatum L.) promoters for green tissue-specific expression of the MYB4 transcription factor for reduced-recalcitrance transgenic switchgrass

Abstract

BackgroundGenetic engineering of switchgrass (Panicum virgatum L.) for reduced cell wall recalcitrance and improved biofuel production has been a long pursued goal. Up to now, constitutive promoters have been used to direct the expression of cell wall biosynthesis genes toward attaining that goal. While generally sufficient to gauge a transgene’s effects in the heterologous host, constitutive overexpression often leads to undesirable plant phenotypic effects. Green tissue-specific promoters from switchgrass are potentially valuable to directly alter cell wall traits exclusively in harvestable aboveground biomass while not changing root phenotypes.ResultsWe identified and functionally characterized three switchgrass green tissue-specific promoters and assessed marker gene expression patterns and intensity in stably transformed rice (Oryza sativa L.), and then used them to direct the expression of the switchgrass MYB4 (PvMYB4) transcription factor gene in transgenic switchgrass to endow reduced recalcitrance in aboveground biomass. These promoters correspond to photosynthesis-related light-harvesting complex II chlorophyll-a/b binding gene (PvLhcb), phosphoenolpyruvate carboxylase (PvPEPC), and the photosystem II 10 kDa R subunit (PvPsbR). Real-time RT-PCR analysis detected their strong expression in the aboveground tissues including leaf blades, leaf sheaths, internodes, inflorescences, and nodes of switchgrass, which was tightly up-regulated by light. Stable transgenic rice expressing the GUS reporter under the control of each promoter (756–2005 bp in length) further confirmed their strong expression patterns in leaves and stems. With the exception of the serial promoter deletions of PvLhcb, all GUS marker patterns under the control of each 5′-end serial promoter deletion were not different from that conveyed by their respective promoters. All of the shortest promoter fragments (199–275 bp in length) conveyed strong green tissue-specific GUS expression in transgenic rice. PvMYB4 is a master repressor of lignin biosynthesis. The green tissue-specific expression of PvMYB4 via each promoter in transgenic switchgrass led to significant gains in saccharification efficiency, decreased lignin, and decreased S/G lignin ratios. In contrast to constitutive overexpression of PvMYB4, which negatively impacts switchgrass root growth, plant growth was not compromised in green tissue-expressed PvMYB4 switchgrass plants in the current study.ConclusionsEach of the newly described green tissue-specific promoters from switchgrass has utility to change cell wall biosynthesis exclusively in aboveground harvestable biomass without altering root systems. The truncated green tissue promoters are very short and should be useful for targeted expression in a number of monocots to improve shoot traits while restricting gene expression from roots. Green tissue-specific expression of PvMYB4 is an effective strategy for improvement of transgenic feedstocks.Electronic supplementary materialThe online version of this article (10.1186/s13068-018-...

McMYB4 improves temperature adaptation by regulating phenylpropanoid metabolism and hormone signaling in apple

“…In maize, ZmMYB31 downregulated the expression of genes involved in monolignol biosynthesis to significantly reduce the lignin content and increase cell wall degradability 33 . In addition, PvMYB4 acts as a master repressor of lignin biosynthesis by decreasing the lignin content and S/G lignin ratio in switchgrass plants 34 . However, MdUGT88F1-mediated phloridzin biosynthesis could increase the levels of lignin and cell wall polysaccharides in apple trees, resulting in increases in internode length and stem and adventitious root numbers and improved growth 35 .…”

“…The samples were then shaken every 6 h during incubation at 4 °C in the dark for 72 h, and the samples were filtered and analyzed by HPLC 6 . The lignin content was measured as described previously and is expressed as OD 280 μg·g −1 (DW) 34 . The contents of endogenous IAA/BR were detected by enzyme-linked immunoassay.…”

Engineering grass biomass for sustainable and enhanced bioethanol production

“…Although CaMV 35S promoters have been implemented for the same, a comprised plant health serves as a major drawback for the same. In a recent study, Liu et al (2018) used a tissue-specific promoter (promoters that offer controlled expression of genes to specific tissues or expressions in specific developmental stages) for the expression Fig. 4 Synthetic two-layer genetic circuits for remodelling of grass cell wall for bioethanol production of PvMYB4 transcription factor gene in switchgrass, which provided a less recalcitrant nature to aboveground biomass of the transgenic switch grass.…”

CsMYB4a from Camellia sinensis Regulates the Auxin Signaling Pathway by Interacting with CsIAA4

“…MYB4 overexpression inhibits the transcriptional expression of phenylpropane pathway genes to reduce the lignin content but also causes growth inhibition, which is called lignin modification-induced dwarfism (LMID) (Panda et al, 2020). In a previous study, to promote the growth of MYB4 transgenic plants, its promoter was replaced, so that PvMYB4 was only overexpressed in green tissues, but not in the roots; this does not affect the growth of the above-ground parts of plants, while reducing their lignin content (Liu et al, 2018).…”