Clint Chapple

Active Mentor - currently hosting PULSe students for laboratory rotations and recruiting PULSe students into the laboratory; serves on preliminary exam committees

Current Research Interests:

The role of the Mediator complex in the regulation of carbon allocation to phenylpropanoid metabolism

The sun is the principle source of energy for our planet, and photosynthesis is the primary mechanism by which that energy is captured and stored in the form of reduced carbon. An outcome of these biochemical events is that plants represent a quantitatively important, sustainable, and carbon-neutral source of energy for humans. In order to maximize the utility of plants for this purpose, it is important that we gain control of the processes associated with energy capture and storage, including the molecular mechanisms that allocate fixed carbon to the myriad biochemical pathways in plants. One of the most significant of these is the phenylpropanoid biosynthetic pathway which leads to the deposition of lignin. Lignin is a cross-linked phenolic polymer that makes the cell walls of specialized plant cells more rigid. Its synthesis represents the single largest metabolic sink for phenylalanine in the biosphere and as such represents a huge metabolic commitment for plant metabolism. Lignin is also a significant barrier to the use of crops for livestock feed, pulp and paper production, and to the generation of cellulosic biofuels. Our objective is to push forward our basic understanding of lignin biosynthesis while simultaneously adding to our ability to engineer plant metabolism so that it can be modified for the improvement of agriculture.

Although the enzymes of lignin biosynthesis have now been identified, we know relatively little about how their expression is regulated. Several relevant transcription factors have been isolated, but it is unclear how their expression and activity dictate or contribute to the allocation of photosynthate to lignin as opposed to other plant components such as cellulose, starch, or any other sinks for reduced carbon.

We are in a unique position to explore how the amount of lignin in a plant is controlled because we have identified two novel plant-specific proteins (REF4 and RFR1) that are components of Mediator, a large multi-protein complex that facilitates interactions between DNA-bound transcription factors and RNA polymerase II to activate or repress the expression of downstream genes. Mutants of Arabidopsis that lack REF4 and RFR1 are viable and show little in the way of developmental changes, making them a tractable system in which to examine the function of Mediator. Of particular relevance to this project is that these mutants accumulate more phenylpropanoid end products including lignin. Plants carrying a mutant dominant form of REF4 show the opposite phenotype. Thus, REF4 and RFR1 appear to be components of a system that determines the amount of carbon allocated to the phenylpropanoid biosynthetic pathway. Considering that over 108 gigatons of lignin are synthesized annually in the biosphere, these proteins are important players in the global carbon cycle and represent important new opportunities for the manipulation of lignin synthesis in plants.

Exploring novel metabolic pathways in Arabidopsis

We have discovered a group of metabolites in Arabidopsis which we have named arabidopyrones (APs). APs are previously undiscovered molecules, the synthesis of which requires the activity of a ring-cleavage dioxygenase, a member of a class of enzymes of mostly unknown function that is conserved across the plant kingdom and beyond. By LC-MS and NMR we have shown APs to be substituted pyrones, the most abundant of which we have named arabidopyl alcohol. The structures of these molecules is highly reminiscent of compounds such as stizolobic and stizolobinic acids, as well as betalamic acid, a component of well-known pigments from beet, Portulaca and various cacti. These tyrosine-derived molecules are all 6-membered N- or O-containing heterocycles bearing substituted 2- or 3-carbon side chains. A common feature of the synthesis of these compounds is that they are derived by recyclization of extra-diol cleavage products of dihydroxyphenylalanine (DOPA). We have found that the only ring cleavage dioxygenase known to be encoded by the Arabidopsis genome (AtLigB) is required for arabidopyrone synthesis, presumably for cleavage of a dihydroxy-substituted precursor.

The fact that ring cleavage dioxygenases have been conserved over 400 million years of plant evolution suggests that they are likely to play an important and conserved role in plant biochemistry. As a result, we believe that the activity of this class of proteins plays a more widespread and fundamental role in plant metabolism that remains to be discovered and that AtLigB has been recruited to serve a specialized role in AP biosynthesis in Arabidopsis. Dissection of AP synthesis in Arabidopsis using genetic, molecular and biochemical tools will shed light on the role(s) of this group of highly specialized catalysts in plants.

Selected Publications:

Dolan WL, Dilkes BP, Stout JM, Bonawitz ND, Chapple C (2017) Mediator Complex Subunits MED2, MED5, MED16, and MED23 genetically interact in the regulation of phenylpropanoid biosynthesis. Plant Cell 29: 3269-3285.

Li Y, Kim JI, Pysh L, Chapple C (2015) Four isoforms of Arabidopsis thaliana 4-coumarate: CoA ligase (4CL) have overlapping yet distinct roles in phenylpropanoid metabolism. Plant Physiol. 169: 2409-2421.

Anderson NA, Bonawitz ND, Nyffeler KE, Chapple C. (2015) Loss of FERULATE 5-HYDROXYLASE leads to Mediator-dependent inhibition of soluble phenylpropanoid biosynthesis in Arabidopsis. Plant Physiol, 169: 1557-1567

Anderson NA, Tobimatsu Y, Ciesielski PN, Ximenes E, Ralph J, Donohoe BS, Ladisch M, Chapple C. (2015) Manipulation of guaiacyl andsyringyl monomer biosynthesis in an Arabidopsis cinnamyl alcohol dehydrogenase mutant results in atypical lignin biosynthesis and modified cell wall structure. Plant Cell 27: 2195-2209.

Kim JI, Dolan WL, Anderson NA, Chapple C. (2015) Indole glucosinolate biosynthesis limits phenylpropanoid accumulation in Arabidopsis thaliana. Plant Cell 27: 1529-1546.

Li X, Svedin E, Mo H, Atwell S, Dilkes BP, and Chapple C. (2014) Exploiting natural variation of secondary metabolism identifies a gene controlling the glycosylation diversity of dihydroxybenzoic acids in Arabidopsis thaliana. Genetics 198: 1267-1276

Bonawitz ND, Kim JI, Tobimatsu Y, Ciesielski PN, Anderson NA, Ximenes E, Maeda J, Ralph J, Donohoe BS, Ladisch M, Chapple C. (2014) Disruption of Mediator rescues the stunted growth of a lignin-deficient Arabidopsis mutant. Nature 509: 376-380.