The Synthetic Biology of N2-Fixing Cyanobacteria for Photosynthetic Terpenoid Production

In the last few decades, concerns over global climate change, energy security, and environmental pollution have been rising. To overcome these challenges, the concept of “-nth generation” biofuels has emerged as a strategy to convert solar radiation into fuels and bulk industrial chemicals for societal use, while decreasing our consumption of nonrenewable energy sources. Nitrogen-fixing cyanobacteria hold a distinct advantage in biofuel production over plants, given their ability to convert sunlight, air (CO2 and N2), and mineralized water to energy-dense carbon molecules, as well as fix atmospheric nitrogen gas into ammonia for metabolism. Engineered cyanobacteria with re-wired metabolic pathways have recently been designed through synthetic biology, and they possess the ability to synthesize new chemicals and biofuels, which are secreted from their cells. Terpenoids constitute one of the largest classes of organic molecules on Earth, and are attractive candidates as a fourth generation biofuel and industrial chemical. In cyanobacteria, the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway is responsible for building essential metabolites involved in photosynthesis, as well as precursors for terpenoid biosynthesis. This dissertation encompasses research focused on redirecting MEP flux in the nitrogen-fixing cyanobacterium Anabaena sp. PCC 7120 to engineered terpenoid sinks, namely, linalool (C10H18O) and farnesene (C15H24). Chapter 1 is a review of literature in the field of biofuels and cyanobacteria, and chapter 2 is an introduction/list of objectives for the research in this dissertation. In chapter 3, we present the genetic engineering of Anabaena to synthesize farnesene by expressing a plant farnesene synthase. In chapter 4, we present the genetic engineering of Anabaena to synthesize linalool during N2-fixation, and increased linalool production is accomplished by the over-expression of three ratelimiting enzymes in the MEP pathway. In chapter 5, we examine the feasibility of a blocking a native carbon reservoir in the cyanobacterium to increase metabolite and energy availability for terpenoid synthesis, as well as physiological aspects of glycogendeficiency in the cyanobacterium during diazotrophic growth. In chapter 6, we focus on introducing a synthetic photorespiratory bypass to reduce photorespiration and increase carbon partitioning towards linalool synthesis

A BioID-Derived Proximity Interactome for SARS-CoV-2 Proteins

The novel coronavirus SARS-CoV-2 is responsible for the ongoing COVID-19 pandemic and has caused a major health and economic burden worldwide. Understanding how SARS-CoV-2 viral proteins behave in host cells can reveal underlying mechanisms of pathogenesis and assist in development of antiviral therapies. Here, the cellular impact of expressing SARS-CoV-2 viral proteins was studied by global proteomic analysis, and proximity biotinylation (BioID) was used to map the SARS-CoV-2 virus-host interactome in human lung cancer-derived cells. Functional enrichment analyses revealed previously reported and unreported cellular pathways that are associated with SARS-CoV-2 proteins. We have established a website to host the proteomic data to allow for public access and continued analysis of host-viral protein associations and whole-cell proteomes of cells expressing the viral-BioID fusion proteins. Furthermore, we identified 66 high-confidence interactions by comparing this study with previous reports, providing a strong foundation for future follow-up studies. Finally, we cross-referenced candidate interactors with the CLUE drug library to identify potential therapeutics for drug-repurposing efforts. Collectively, these studies provide a valuable resource to uncover novel SARS-CoV-2 biology and inform development of antivirals

Via Egnatia after Egnatius: Imperial policy and inter-regional contacts

The Via Egnatia, which linked Dyrrachium to Kypsela and ultimately to Byzantium/ Constantinople, was the first Roman highway to be built east of the Adriatic. The studies published so far on this important road are devoted almost exclusively to its military importance, particularly during the Roman Republic. This author's goal instead was to assess the importance of the Egnatia at apolitical, social, and cultural level, by examining written sources (literary and epigraphical) and material remains. The article looks into the policy of Roman emperors regarding the Egnatia, and the role of the Via as a factor of commercial, social, and cultural interaction between the Italian peninsula and the Greek world, as well as among the cities and regions that it crossed. It also shows the contribution of the Egnatia to the spectacular development of certain cities and the parallel weakening of others, together with its impact upon the rural landscape