The hyperthermophilic archaeon Pyrococcus furiosus (Pf) grows optimally at 100°C by fermenting peptides and sugars. It also reduces elemental sulfur to hydrogen sulfide. From Pf we are purifying and characterizing a range of metal-containing, oxidoreductase-type enzymes and redox proteins that are involved in unusual catabolic pathways. In addition, all ORFs in the Pf genome (1.9 Mb) are being cloned and expressed in an NIH-funded structural genomics initiative with the goal of obtaining 3D structures on all Pf proteins. The function of all Pf ORFs are being assessed using DNA microarrays and proteomic approaches in conjunction with metabolic and physiological analyses.
Dr. Carlson's research is directed toward characterizing the molecular basis for the interaction between a bacterium and a plant or animal host cell. One system under examination is the nitrogen-fixing symbiotic infection of legumes by rhizobia. These nitrogen-fixing soil bacteria contain genes that are activated by flavonoid molecules produced by the host plant. These genes encode enzymes which synthesize a glycolipid which is an acylated chitin oligosaccharide. In most cases, the glycolipids produced by each Rhizobium species are structurally modified which results in their ability to interact only with a specific legume plant. This molecular recognition process results in the stimulation of cell division in the legume root causing a nodule to form. The cells in this nodule are invaded by the rhizobia and are where nitrogenase is produced which reduces dinitrogen to ammonia. Other molecules on the surface of rhizobia that are required for the invasion of the root nodule cells by these bacteria are the outer membrane capsular and lipopolysaccharides. Specific structural changes occur in these molecules in response to the host plant which are crucial for infection. These structural changes, and the genes that are responsible for them, are presently under investgation. Dr. Carlson also has projects directed toward characterizing the role that bacterial lipopolysaccharides and lipooligosaccharides play in determining the pathogenicity of such organisms as Salmonella enteritidis, Neiserria meningiditis, and Hemophilus influenzae. Both the plant symbiont and animal pathogen work are being done in collaboration with several research groups in other universities. Dr. Carlson's research is currently funded by two grants from the USDA, one grant from the NSF, and one grant from the NIH.
Darvill's research focuses on structurally characterizing the five major noncellulosic carbohydrates of plant primary cell walls homogalacturonan, rhamnogalacturonan I and II, xyloglucan, and glucuronoarabinoxylan. Plant primary cell walls control the rate and direction of cell growth that determine ultimately the shapes of cells, tissues, and organs; they form a barrier to pathogens, are the source of oligosaccharins that elicit plant defense responses to pathogens, and participate in controlling plant growth and development. New analytical techniques are continually developed to isolate and determine the complicated structures and functions of these molecules. Interactions between wall-matrix polysaccharides is being characterized by examining the cell-, tissue-, and species-dependent expression of cell wall epitopes using well-characterized monoclonal antibodies.
Our research is directed toward an improved understanding of the biochemical and molecular genetic mechanisms that control plant growth and development, particularly with respect to vascular tissues and lignocellulose formation. In this effort, we use a suite of genomic approaches based on high-throughput DNA sequencing and microarray platforms. We are heavily involved in an international effort to create a reference genome for conifers, and use loblolly pine (Pinus taeda) for this work. At the level of specific genes and pathways we are very interested in multicopper oxidases, such as laccase, and in the ethylene biosynthetic pathway.
Pectin is a family of complex polysaccharides present in all plant primary cell walls. Pectin plays multiple roles in plant growth, development, and defense responses; in part through contributing to cell wall strength, wall ion exchange and sieving properties, cell-cell adhesion, and cell-cell communication. Pectin is a food fiber and a commercial gelling agent that has beneficial effects on human health. Our long term goal is to decipher how the 53 distinct enzyme activities required for pectin synthesis interact to synthesize pectin and to modify pectin synthesis in order to study pectin function. Towards this goal we are purifying, cloning, and characterizing the biosynthetic enzymes; many of which are Golgi localized and membrane bound enzymes. Current emphasis is on the galacturonosyltransferase and the methyltransferase that synthesize the pectic polysaccharide homogalacturonan.
RNA-guided invader defense in prokaryotes: Archaea and bacteria (both pathogenic and beneficial) are constantly attacked and destroyed by viruses and other genome invaders. We are working to delineate a series of newly-identified RNA-mediated immune systems that protect prokaryotes from viruses and other invaders - the CRISPR-Cas systems. This exciting research is leading to new ways to strengthen beneficial microorganisms that produce food, pharmaceuticals and biofuels, to combat disease-causing bacteria, and to prevent the spread of antibiotic resistance.
The "primary" cell wall, which surrounds growing plant cells, plays a key role in plant development. One of its most important functions is to control the rate and orientation of cell expansion. Polysaccharide networks in the wall expand by gradually yielding under osmotic stress, allowing the cell to grow in a controlled, oriented fashion. This process determines the morphology of each cell, which ultimately determines the shape of the entire plant. Research in my laboratory is aimed at characterizing the molecular dynamics and topology that lead to the assembly and controlled expansion of the cell wall.