Jason A. Carlyon, Ph.D.

Assistant Professor

Selected publications

A. phagocytophilum adhesins that facilitate attachment to neutrophils. The initial host-pathogen interaction preceding bacterial colonization is mediated by bacterial factors called adhesins. These surface proteins have specific structural requirements for targeting their ligands, which results in colonization being restricted to certain cell populations that display the optimal receptors. The remarkable tropism of A. phagocytophilum for neutrophils is attributable to its usage of P-selectin glycoprotein ligand-1 (PSGL-1) and sialylated and alpha 1,3-fucosylated glycans on neutrophil surfaces for cellular adhesion. We, in collaboration with Dr. Rodger P. McEver of the Oklahoma Medical Research Foundation and Dr. Richard D. Cummings of the University of Oklahoma Department of Biochemistry and Molecular Biology, further defined the A. phagocytophilum cytoadherence mechanism by identifying two key molecular features of PSGL-1 to which the organism binds: (1) a primary amino acid sequence found in the N-terminus of human PSGL-1 and (2) sialyl Lewis x (sLe x), a sialylated and fucosylated tetrasaccharide that caps PSGL-1 and other selectin ligands. The bacterial protein(s) that mediate these interactions are unknown, which represents a considerable gap in our knowledge. We hypothesize that A. phagocytophilum adherence to neutrophils is dependent on one or more adhesins that target PSGL-1 N-terminal peptide and sLe x. Identifying the cognate adhesins will be integral to developing strategies for disrupting the interaction of A. phagocytophilum with the surface of its host cell. The objectives of this project are to identify and characterize the A. phagocytophilum adhesin(s) that mediate cytoadherence to human neutrophils. This will involve: (1) isolating putative adhesins based on their affinities for PSGL-1 and sLe x; (2) identifying, cloning, and expressing them using proteomic and molecular methods; (3) functionally characterizing the adhesin candidates using glycoconjugate and cellular binding assays; and (4) assessing whether disrupting their adhesion activities can inhibit infection in vitro and in vivo. Accomplishing these goals will further our knowledge of A. phagocytophilum pathogenesis and bacterial adhesion strategies. It will identify targets for potentially treating or preventing HGA and may foster development of new pharmacologic inhibitors of cellular adhesion events associated with inflammatory disorders.

A. phagocytophilum invasion and intracellular survival mechanisms. Neutrophils are the first line of defense against invading pathogens. One of the primary means by which neutrophils destroy phagocytosed bacteria is dependent on superoxide anion (O 2 - ), which is produced by the tightly controlled, rapidly activatable NADPH oxidase complex. In resting neutrophils, the oxidase components are unassembled. Upon stimulation, the components localize to the cell membrane and, at the expense of NADPH, generate O 2 - either outside the cell or within the lumen of the phagosome containing an ingested bacterium. Research by our group and others have demonstrated that A. phagocytophilum utilizes a multi-tiered approach to avoid and subvert neutrophil oxidative killing. During host cell binding and invasion, the bacterium uses an undefined mechanism to rapidly detoxify O 2 - in its local environment. Once inside, A. phagocytophilum resides within a protective vacuole that minimally excludes fusion with specific granules carrying cytochrome b 558, an integral component of NADPH oxidase. The mechanism by which this is accomplished and whether other oxidase components are excluded are unknown. Upon establishing residence, the organism interferes with host transcription of gp91 phox and rac2, which encode two key oxidase components, thereby rendering its host neutrophil oxidase-deficient. Many exciting projects have stemmed from this work. We are interested in defining the means by which A. phagocytophilum establishes residence within its host cell and excludes cytochrome b 558 and other host cellular components from its parasitophorous vacuole. This will involve (1) proteomic dissection of the A. phagocytophilum vacuole to identify bacterial proteins that may participate in either construction of the vacuole or inhibiting intracellular trafficking of host cellular components to the vacuole. This will also incorporate (2) differential expression analyses to identify A. phagocytophilum genes that are up-regulated upon bacterial interaction with PSGL-1, association with the host cell membrane, and during the initial stages of infection. The identified genes and their encoded proteins will be assessed for their potential roles in invasion and in conditioning the host cell for bacterial residence.

The first case of HGA was documented in 1994, making this a young field ripe for investigation. Because it is an intracellular pathogen, studying A. phagocytophilum entails a multidisciplinary approach involving aspects of microbiology, cell and molecular biology, immunology, and biochemistry. Deciphering the adherence, invasion, and intracellular survival strategies of this unique organism will not only further our understanding of A. phagocytophilum pathogenesis, but also offers a novel approach to comprehending neutrophil biology and defense mechanisms. There are many exciting research projects available in my lab. If interested, please do not hesitate to contact me.