Increasing the Value of Bioenergy Grasses—Expressing Engineered Traits in the Right Place at the Right Time
Authors:
Jie Fu1,2*(jief3@illinois.edu), Brian McKinley3,4, Brandon James5, William Chrisler6, Lye Meng Markillie6, Matthew J Gaffrey6, Hugh D Mitchell6, Galya Orr6, Kankshita Swaminathan2,5*, John Mullet3,4, and Amy Marshall-Colon1,2
Institutions:
1University of Illinois Urbana–Champaign; 2Center for Advanced Bioenergy and Bioproducts Innovation; 3Texas A&M University; 4Great Lakes Bioenergy Resource Center (GLBRC); 5HudsonAlpha Institute for Biotechnology; and 6Pacific Northwest National Laboratory
URLs:
Goals
The overarching goal of the Center for Advanced Bioenergy and Bioproducts Innovation (CABBI) Feedstock Production Theme research is to deliver resilient, highly productive grasses that contain large amounts of lipids. Researchers have made significant advances in engineering production of oils, specialty fatty acids, and other organic compounds in vegetative tissues of these grasses, as well as increasing biomass yield, resource use efficiency, and environmental resilience. A specific challenge researchers are still working on is targeting expression of engineered traits in the right place at the right time, paving the way for CABBI crops that produce oil in stem storage tissues at the end of the season.
Abstract
Overview: To contribute to the development of the bioeconomy, feedstocks need to be improved to be economically and environmentally sustainable for both processor and farmer. CABBI strives to do this by engineering carbon allocation to produce oils, specialty fatty acids, and other organic compounds, as well as by increasing biomass yield, resource use efficiency, and environmental resilience. Because such improvements require coordinated changes in multiple plant traits, researchers are working simultaneously on trait improvements and how best to combine and implement them to get the traits in the right place at the right time. In the high biomass C4 grass feedstocks—sorghum, miscanthus and sugarcane—the right place for value-added products is stems. As the right time for different traits varies, researchers are performing analyses across multiple key developmental time points.
Spatial atlas of the sorghum stem: Bioenergy sorghum’s 4-5m stems account for ~80% of the harvested biomass. Stems accumulate high levels of sucrose that could be used to synthesize bioproducts if information about stem cell-type gene expression and regulation was available to enable engineering. To obtain this information, Laser Capture Microdissection (LCM) was used to isolate transcriptome profiles from five major cell types present in vegetative stems of Sorghum bicolor L. Moench cv. Wray. Transcriptome analysis identified genes with cell-type specific and cell-preferred expression patterns that reflect the distinct characteristics and regulatory functions of each cell type. The newly discovered cell type specific genes can be used as markers for downstream analyses, such as single cell transcriptomics. Analysis of cell-type specific gene regulatory networks (GRNs) revealed that 1) different biological functions distinguish vascular and non-vascular cell types; 2) distinct transcription factor families regulate the cell type specific expression of genes; and 3) cell type specific transcription factors have both direct and indirect modes of regulation to modulate the expression of cell type specific genes. The team used the LCM data to gain insights into stem secondary cell wall (SCW) networks. By combining the spatial resolution of the LCM-derived stem cell-type specific transcriptome with a stem developmental profile of SCW formation, researchers uncovered 1) the previously unknown spatial expression of key SCW genes across sorghum stem cell types, 2) cell type specific SCW regulatory networks and network motifs, and 3) potential regulators that repress SCW formation in pith parenchyma cells. The cell-type transcriptomic dataset provides a valuable source of information about the function of sorghum stems and GRNs that will enable the engineering of bioenergy sorghum stems.
Future directions: The spatial transcriptomes provide rich information about steady-state gene expression and identified cell-type specific hub genes that likely play key regulatory roles in signaling within each cell type. However, the products of gene expression, the proteome and metabolome, better reflect macro-level phenotypes because they strengthen the link between gene expression and gene function. A continuing collaboration among CABBI, GLBRC, and Environmental Molecular Sciences Laboratory (EMSL) will expand the sorghum stem molecular atlas by incorporating spatial proteomic and metabolomic data into the existing GRNs, which will improve network predictions by more directly linking genes to stem phenotypes. Likewise, researchers will expand the network analysis to include spatial transcriptomes from two additional developmental time points (onset of anthesis and post-anthesis). These combined analyses will reveal the spatio-temporal dynamics of stem metabolic and signaling networks. This deep understanding of these spatio-temporal dynamics is critical to being able to engineer stems and redirect bioproducts to the right place at the right time.
Connections across Feedstocks: An actionable outcome of the above analyses is the identification of specific promoter elements that drive cell type specific expression at different points in development. Such molecular tools provide a direct path to engineer sorghum stems to accumulate high-value bioproducts of interest to CABBI and its sister Bioenergy Research Centers (BRCs) and decrease the burden on conversion groups in the biofuel industry. Hence, this is a cross-BRC priority. With the close relation between sorghum, sugarcane, and miscanthus, the team expects that these findings will provide insights into stem-specific expression across the feedstocks of interest. The team also expects many of the findings and methodologies from this study to be useful to researchers interested in engineering other grasses of interest, such as switchgrass and maize. Combining this knowledge with CABBI advances in accumulating oil in sugarcane and sorghum, improvements in water use efficiency and photosynthesis, and breeding to identify plants with higher yields and greater geographic range will generate feedstocks that can be a foundation for a strong bioeconomy.
Funding Information
This work was funded by the DOE Center for Advanced Bioenergy and Bioproducts Innovation (U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research under Award Number DE-C0018420). Any opinions, findings, and conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the views of the U.S. Department of Energy.