Many pathogens encode proteases that serve to antagonize the host immune system. In particular, viruses with a positive-sense single-stranded RNA genome ((+)ssRNA), including picornaviruses, flaviviruses, and coronaviruses, encode proteases that are not only required for processing the viral polyprotein into functional units but also manipulate crucial host cellular processes through their proteolytic activity. Because these proteases must coordinate both the cleavage of numerous polyprotein sites and subversion of host immunity, evolution of viral proteases is expected to be highly constrained. Despite this strong evolutionary constraint, these viral proteins and host immune factors are engaged in “evolutionary arms races” that results in diverse protease-host interactions even within closely related species. In some cases, rapid host gene evolution can result in avoidance of cleavage by viral proteases. In other, more recently described cases, hosts can evolve to bait viral proteases into cleaving them using a “tripwire” strategy of immune activation. Such data provide an explanation for why viral polyprotein sites evolve despite such a strong evolutionary constraint and highlight the importance of identifying and characterizing host proteins that are targeted by viral proteases. Moreover, such an evolutionary model provides insight into the changes in molecular functions between viral proteases and host factors, and underscores the role of viral proteases in viral host range, zoonosis and host immune gene evolution. Here, I describe a combined computational and functional approach that guide the discovery of new host targets of viral proteases, including the characterization of two new host innate immunity tripwires that trigger inflammation in response to viral protease activity.

ABSTRACT OF THE THESIS

The LysE Superfamily of Transport Proteins Involved in Cell Physiology and Pathogenesis

by

Brian Vay Tsu

Master of Science in Biology

University of California, San Diego 2015

Professor Milton Saier, Chair

The LysE superfamily consists of transmembrane transport proteins that catalyze export of amino acids, lipids and heavy metal ions. Statistical means were used to show that it includes newly identified families including transporters specific for (1) tellurium, (2) iron/lead, (3) manganese, (4) calcium, (5) nickel/cobalt, (6) amino acids, and (7) peptidoglycolipids as well as (8) one family of transmembrane electron carriers. Internal

repeats and conserved motifs were identified, and multiple alignments, phylogenetic trees and average hydropathy, amphipathicity and similarity plots provided evidence that all members of the superfamily derived from a single common 3-TMS precursor peptide via intragenic duplication. Their common origin implies that they share common structural, mechanistic and functional attributes. The transporters of this superfamily play important roles in ionic homeostasis, cell envelope assembly, and protection from excessive cytoplasmic heavy metal/metabolite concentrations. They thus influence the physiology and pathogenesis of numerous microbes, being potential targets of drug action.