Theodore Clark, PhD
Department of Microbiology and Immunology
Professor of Parasitology and Immunology
Department of Microbiology and Immunology
Cornell University College of Veterinary Medicine
C5 163 VMC
Ithaca, NY 14853
Our laboratory focuses on the biology of host-pathogen interactions, and, more broadly, the control of infectious disease. We have a long-standing interest in Ichthyophthirius multifiliis, a parasitic ciliate that causes “white-spot” disease, or “Ich” in freshwater fish. Ichthyophthirius is widely distributed in nature and has significant impact on commercial aquaculture worldwide. At the same time, it serves as a useful model for the study of host-pathogen interactions in lower vertebrates where adaptive immunity first evolved. In parallel with our work on Ichthyopthirius, we study the free-living non-pathogenic ciliate, Tetrahymena thermophila, as a surrogate for understanding the biology of I. multifiliis, and as an alternative platform for the production of recombinant subunit vaccines. On the host side, we are interested in questions surrounding the evolution of adaptive immunity and have identified and characterized functional homologs of mammalian dendritic cells in rainbow trout. Finally, we made the recent discovery that viral envelope proteins responsible for the entry of large numbers of viruses (dengue, Zika, West Nile, etc.) into host cells bear a striking resemblance to HAP2, a gamete fusion protein that mediates fertilization in a wide range of taxa. The implications of this finding for the emergence of eukaryotic sex, and possible evolutionary origins of viral fusogens are currently being explored.
The Biology and Control of Ichthyophthirius. Aquaculture is the fastest growing sector of agribusiness worldwide, with farm-raised fish accounting for ~30% of the world’s annual catch. Because of its broad host range and geographic distribution, Ichthyophthirius multifiliis is among the most important protozoan pathogens of farm-raised fish. Our research is directed towards the development of vaccines and small molecule drugs for the treatment and prevention of “white-spot”. Efforts at vaccine development have focused on class of abundant GPI-anchored surface proteins known as immobilization antigens (i-antigens) that are the primary targets of the host immune response. Antibodies against these proteins immobilize the parasite in vitro and trigger a novel evasion strategy in vivo in which parasites exit fish prematurely to evade the host immune response. Along with premature exit, recent studies show that surface antigen clustering following antibody binding can induce extrusion of large numbers of mitochondria from both Ichthyophthirius and from its free-living cousin, Tetrahymena thermophila. The biological significance of this phenomenon is now being studied. While the i-antigens remain obvious vaccine candidates, we have recently completed large-scale transcriptional profiling studies along with sequencing of the I. multifiliis macronuclear genome with the hope of identifying additional vaccine and drug targets for the prevention and treatment of Ich. Along with its practical importance, the genome sequence provides a powerful tool for understanding the biology of this organism, including as yet unidentified stages of the parasite life cycle.
Tetrahymena as a Vaccine Production Platform. Long a model for basic research, Tetrahymena offers a number of salient features for recombinant protein production including rapid scalable growth; eukaryotic machinery for protein folding and post-translational modification; a metabolism geared towards membrane protein production; and, the ability to introduce transgenes at very high copy number leading to robust protein expression. This novel technology platform has proven to be extremely useful for the production of otherwise difficult to express eukaryotic membrane proteins such as voltage-dependent ion channels. We are currently using Tetrahymymena for large-scale expression of candidate vaccine antigens from influenza virus, Plasmodium falciparum (a causative agent of malaria) and Ichthyophthirius multifiliis.
Evolution of Adaptive Immunity. Dendritic cells (DCs) are specialized antigen-presenting cells that bridge innate and adaptive immunity. Although critical to the development of T-cell responses in mammals, it is not known whether DCs co-evolved with adaptive immunity, or if antigen presentation relied on a fundamentally different cell type early in vertebrate evolution. We are exploring this question in rainbow trout (Oncorhynchus mykiss) as a representative of teleost fish, among the earliest extant species to have evolved acquired immune responses. Adapting protocols for generating DCs in mammals, we have developed methods to culture highly motile non-adherent DC-like cells from trout that have irregular membrane processes and express surface MHCII. In mixed leukocyte reactions these cells have been shown to be superior to either macrophages or B-cells in inducing proliferative responses by trout T-cells. In addition, they have many of the hallmarks of mammalian dendritic cells including tree-like morphology; the expression of DCs markers; the ability to phagocytose small particles; activation by toll-like receptor ligands; and, the ability to migrate in vivo. Although DCs have recently been identified in zebrafish, the ability to culture large numbers of these cells from trout hematopoietic tissues should provide a useful approach to the study of teleost DC function at the cellular and biochemical levels.
HAP2 and the origins of eukaryotic sex. HAP2 is a highly conserved transmembrane protein that can be traced to the basal lineages all major branches the eukaryotic tree of life. We and others have recently demonstrated that HAP2 is an ancient gamete fusogen mediating sperm-egg fusion in sexually dichotomous species, and mating-type fusion in Tetrahymena, which bypasses the production of “male” and “female” gametes altogether and instead produces stationary (“female”) and migratory (“male”) pronuclei that are exchanged between mating cells. Of particular interest is the fact that HAP2 shares all the major structural features of class II viral fusion proteins. Thus, HAP2 may have arisen with a virus (through lateral gene transfer) and been instrumental in the origins of eukaryotic sex, or it may have been repurposed by an early eukaryotic virus (or parasitic DNA element) for cell-to-cell spread. In either case we are studying the structural requirements for HAP2 function in Tetrahymena and looking for structural analogs that may catalyze sperm-egg fusion in taxa that no longer contain the HAP2 gene (most importantly humans).
PhD (SUNY at Stony Brook)
Dr. Clark joined the Department of Microbiology and Immunology in 1996 and is currently interim Department Chair. Dr. Clark received his BS from Columbia University, his PhD from the State University of New York at Stony Brook and was a Postdoctoral Fellow in the Department of Biology at Yale University before moving to the University of Georgia as a Research Associate. Over the years, his research has been supported by grants from the National Institutes of Health, the National Science Foundation, the Bill & Melinda Gates Foundation, the Department of Defense, and the U.S. Department of Agriculture. He is a member of the Aquatic Animal Medicine Program in the College of Veterinary Medicine, and founded Tetragenetics Inc., an early stage biotechnology company now in Arlington, Massachusetts.
Clark, T. (2018). HAP2/GCS-1: Mounting evidence of our true biological EVE? PLoS Biol. doi: 10.1371/journal.pbio.3000007. PMID:30125288.
Bednenko J, Harriman R, Mariën L, Nguyen HM, Agrawal A, Papoyan A, Bisharyan Y, Cardarelli J, Cassidy-Hanley D, Clark T, Pedersen D, Abdiche Y, Harriman W, van der Woning B, de Haard H, Collarini E, Wulff H, Colussi P. (2018). A multiplatform strategy for the discovery of conventional monoclonal antibodies that inhibit the voltage-gated potassium channel Kv1.3. MAbs. doi: 10.1080/19420862.2018.1445451. [Epub ahead of print].
Pinello JF, Lai AL, Millet JK, Cassidy-Hanley D, Freed JH, Clark TG. (2017). Structure-Function Studies Link Class II Viral Fusogens with the Ancestral Gamete Fusion Protein HAP2. Curr Biol. 27(5):651-660. doi: 10.1016/j.cub.2017.01.049.
Zaila KE, Doak TG, Ellerbrock H, Tung CH, Martins ML, Kolbin D, Yao MC, Cassidy-Hanley DM, Clark TG, Chang WJ. (2017). Diversity and universality of endosymbiotic Rickettsia in the fish parasite Ichthyophthirius multifiliis. Front. Microbiol. 2017 Feb 9;8:189. doi: 10.3389/fmicb.2017.00189
Bisharyan Y., Clark T. (2016). Signaling Through GPI-Anchored Surface Antigens in Ciliates. In: Witzany G., Nowacki M. (eds) Biocommunication of Ciliates. Springer, Cham. doi.org/10.1007/978-3-319-32211-7_9
MacColl E, Therkelsen MD, Sherpa T, Ellerbrock H, Johnston LA, Jariwala RH, Chang W, Gurtowski J, Schatz MC, Mozammal Hossain M, Cassidy-Hanley DM, Clark TG, Chang WJ. (2015). Molecular genetic diversity and characterization of conjugation genes in the fish parasite Ichthyophthirius multifiliis. Mol Phylogenet Evol. 86:1-7. doi: 10.1016/j.ympev.2015.02.017.
Cole ES, Cassidy-Hanley D, Fricke Pinello J, Zeng H, Hsueh M, Kolbin D, Ozzello C, Giddings T Jr, Winey M, Clark TG. (2014). Function of the male-gamete-specific fusion protein HAP2 in a seven-sexed ciliate. Curr Biol. 24(18):2168-2173. doi:10.1016/j.cub.2014.07.064.
Bassity E, Clark TG. (2012). Functional identification of dendritic cells in the teleost model, rainbow trout (Oncorhynchus mykiss). PLoS One. 2012;7(3):e33196. doi: 10.1371/journal.pone.0033196.
Awards and Honors