The Lignin Enigma: Turning the Corner
EMSL Project ID
49285
Abstract
Lignins (Nature's second most abundant terrestrial plant substances) provide structural support for plants to stand upright, to conduct water/nutrients, and as barriers to opportunistic pathogens. Lignin properties were pivotal in evolutionary transition of aquatic plants to dry land, without which life as we know could never exist or be sustained. Yet how lignification occurs in planta, including producing distinct lignin compositions/configurations in different cell types remains unresolved. Lignins/lignocellulosic recalcitrance represent major obstacles in producing cost competitive bioenergy and feedstocks/biofuels at scale and cost, and thus limit enhanced biomass utilization. Recalcitrance impacts global C recycling (greenhouse gas accumulation and terrestrial C turnover in soil and aquatic systems, whether re-mineralized or sequestered). Regarding EMSL mission and 2016 science themes, transformative molecular level discoveries on lignification are urgently needed. Yet, understanding lignification proper will positively impact 2016 topics of Biosystems Dynamics and Design (dynamic inter- and intra-cellular processes), Energy Materials and Processes (including biomass degradation, bio-produced fuels and renewable chemicals), and Terrestrial and Subsurface Ecosystems (C environmental cycling).
Our team recently established that the uniform pattern of lignin deposition in a specific cell type (i.e. Casparian bands) is massively disrupted ("misdirected") when dirigent protein (DP) homologs in this cell type undergo loss-of-function. This led to a potentially game-changing hypothesis of a tightly coordinated lignin-forming supramolecular protein complex, consisting of a CASP scaffold, upon which dirigent proteins (DPs), laccases, peroxidases, super oxide dismutases (SOD) and NOXs are organized to enable defined lignin assembly. There are 2 objectives:
1. Dissect lignin-forming complexes. Establish precise nature of interacting proteins with DP's, particularly those providing 1e-- oxidation, cofactors and potential scaffold roles,
2. Establish DP role(s) in how macromolecular lignin formation is orchestrated.
Our highly collaborative team needs access to the outstanding EMSL capabilities and expertise to establish the precise nature of the proposed lignin forming complex(es) and the lignins so formed, not only in Arabidopsis Casparian strips, but also in distinct lignifying biomass cell walls, such as lignified guaiacyl or syringyl cell walls in poplar. With Casparian strip DP mutants and poplar lines in hand, we can uniquely probe, dissect, and define the true lignin-forming apparatus in different cell types and compare/contrast same. Specifically, we wish to dissect the various lignin forming supramolecular protein complexes and define interactions and functions of the corresponding proteins, as well as nature of lignins formed. This requires access to EMSL Laser Capture Microdissection and FACS analysis capabilities to isolate distinct lignifying cell types and then applying EMSL nanoscale proteomics and cryo-EM to define the nature of the overall protein complex(es), as well as their relative abundances and holistic functions in lignification. Isolation protocols will also be applied to study lignins of specific cell wall types using advanced EMSL NMR and MS capabilities (facilitated by 13C labelling of lignins), as well as in determining molecular weights of WT and DP mutant lignins.
Expected Results: We envisage a new paradigm for lignification, this being of critical importance to current BER/EMSL emphases. Expected results:1) Demonstrate existence of lignin polymerization protein complexes in distinct bimass cell wall types, analogous to CS lignin forming endodermal cells; 2) Identify interactions among components in the lignin polymerizing protein complex, and develop initial structural model of the complex; and 3) establish CS lignin in DP knockouts has molecular defects.
Project Details
Project type
Large-Scale EMSL Research
Start Date
2016-10-01
End Date
2018-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Co-Investigator(s)
Team Members
Related Publications
Corbin C., S. Drouet, L. Markulin, D. Auguin, D. Auguin, E. Laine, and L.B. Davin, et al. 2018. "A genome-wide analysis of the flax (Linum usitatissimum L.) dirigent protein family: from gene identification and evolution to differential regulation." Plant Molecular Biology 97, no. 1-2:73-101. PNNL-SA-131411. doi:10.1007/s11103-018-0725-x
Garcellano R.C., S. Moinuddin, R.P. Young, M.E. Bowden, M. Zhou, R.S. Renslow, and Y. Yesiltepe, et al. 2018. "Isolation of Tryptanthrin and Reassessment of Evidence for its Isobaric Isostere Wrightiadione in Plants of the Wrightia Genus." Journal of Natural Products. PNNL-SA-135279. doi:10.1021/acs.jnatprod.8b00567.
Garcellano R.C., S. Moinuddin, R.P. Young, M.E. Bowden, M. Zhou, R.S. Renslow, and Y. Yesiltepe, et al. 2019. "Isolation of Tryptanthrin and Reassessment of Evidence for its Isobaric Isostere Wrightiadione in Plants of the Wrightia Genus." Journal of Natural Products 82, no. 3:440-448. PNNL-SA-135279. doi:10.1021/acs.jnatprod.8b00567