Jan. 29, 2013 — Researchers seeking to improve production of ethanol from woody crops have a new resource in the form of an extensive molecular map of poplar tree proteins, published by a team from the Department of Energy's Oak Ridge National Laboratory.
Putting the Pieces Together: High-performance LC-MS/MS Provides Network-, Pathway-, and Protein-level Perspectives in Populus
High-performance mass spectrometry (MS)-based proteomics enabled the construction of a detailed proteome atlas for Populus, a woody perennial plant model organism. Optimization of experimental procedures and implementation of current state-of-the-art instrumentation afforded the most detailed look into the predicted proteome space of Populus, offering varying proteome perspectives: (1) network-wide, (2) pathway-specific, and (3) protein-level viewpoints. Together, enhanced protein retrieval through a detergent-based lysis approach and maximized peptide sampling via the dual-pressure linear ion trap mass spectrometer (LTQ Velos), have resulted in the identification of 63,056 tryptic peptides. The technological advancements, specifically spectral-acquisition and sequencing speed, afforded the deepest look into the Populus proteome, with peptide abundances spanning 6 orders of magnitude and mapping to ∼25% of the predicted proteome space. In total, tryptic peptides mapped to 11,689 protein assignments across four organ-types: mature (fully expanded, leaf plastichronic index (LPI) 10–12) leaf, young (juvenile, LPI 4–6) leaf, root, and stem. To resolve protein ambiguity, identified proteins were grouped by sequence similarity (≥ 90%), thereby reducing the protein assignments into 7538 protein groups. In addition, this large-scale data set features the first systems-wide survey of protein expression across different Populus organs. As a demonstration of the precision and comprehensiveness of the semiquantitative analysis, we were able to contrast two stages of leaf development, mature versus young leaf. Statistical comparison through ANOVA analysis revealed 1432 protein groups that exhibited statistically significant (p ≤ 0.01) differences in protein abundance. Experimental validation of the metabolic circuitry expected in mature leaf (characterized by photosynthesis and carbon fixation) compared with young leaf (characterized by rapid growth and moderate photosynthetic activities) strongly testifies to the credibility of the approach. Instead of quantitatively comparing a few proteins, a systems view of all the changes associated with a given cellular perturbation could be made.
Populus, a fast-growing perennial tree, holds potential as a bioenergy crop due to its ability to produce large amounts of biomass on non-agricultural land. Now, a study by ORNL scientists with the Department of Energy’s BioEnergy Science Center has provided the most comprehensive look to date at poplar’s proteome, the suite of proteins produced by a plant’s cells. The study is featured on the cover of January’s Molecular & Cellular Proteomics.
“The ability to comprehensively measure genes and proteins helps us understand the range of molecular machinery that a plant uses to do its life functions,” said ORNL’s Robert Hettich. “This can provide the information necessary to modify a metabolic process to do something specific, such as altering the lignin content of a tree to make it better suited for biofuel production. “
The ORNL research team measured more than 11,000 proteins in different parts of poplar, including mature leaves, young leaves, roots and stems. This systematic approach yielded a so-called proteome atlas, which maps out the proteins present in the various tissue types at a given point in time. Lead coauthors Paul Abraham and Richard Giannone describe how the atlas offers a broad overview of the poplar proteome and also the ability to zoom in on specific biological features, such as pathways and individual proteins.
“We tried to provide a zoomable view, like Google maps, so you can look at the system from various perspectives,” Abraham said. “By having these different viewpoints, it makes it easier to mine out the relevant biological information.”
Obtaining and analyzing information about plant proteomes is especially tricky, considering a plant such as poplar can potentially manufacture more than 40,000 different proteins. Unlike an organism’s genome, which is the same for every cell and remains constant, the proteome varies from cell to cell and changes over time as the plant adapts to different environmental conditions.
“The analytical techniques we’ve demonstrated allow us to measure the range of proteins very deeply and specifically, so we can start to figure out, for instance, how the protein machinery in a leaf differs from the ones in the trunk,” Hettich said. “Or we can look at a tree that’s very young versus one that’s very old, thus enabling us to understand how all these proteins are changing as a function of the tree growing older.”
Knowing how plants change and adapt to environmental surroundings by altering their proteins could help bioenergy researchers develop poplar trees better suited to biofuel production.
“It’s the proteins that directly alter the morphology, anatomy, and function of a plant cell,” Abraham said. “If we can identify the proteins that create a favorable trait such as fast growth, then we can incorporate that protein or modify it to develop a superior plant with all favorable traits through transgenics.”
The study’s coauthors are ORNL’s Robert Hettich, Paul Abraham, Richard Giannone, Rachel Adams, Udaya Kalluri and Gerald Tuskan.
BESC is one of three DOE Bioenergy Research Centers established by the DOE’s Office of Science in 2007. The centers support multidisciplinary, multi-institutional research teams pursuing the fundamental scientific breakthroughs needed to make production of cellulosic biofuels, or biofuels from nonfood plant fiber, cost-effective on a national scale. The three centers are coordinated at ORNL, Lawrence Berkeley National Laboratory and the University of Wisconsin-Madison in partnership with Michigan State University.
ORNL is managed by UT-Battelle for the Department of Energy’s Office of Science. DOE’s Office of Science is the single largest supporter of basic research in the physical sciences in the United States, and is working to address some of the most pressing challenges of our time. For more information, please visit science.energy.gov.
Press release and image courtesy of Oak Ridge National Laboratory. You can find the original press release here.