LS² Annual Meeting 2022

Plants, fungi and bacteria produce a wealth of specialized metabolites, which are of great importance from both ecological and clinical perspectives. Due to the accelerated accumulation of omics data, computational methods have become more and more important to identify these molecules and to assess their biological activities. Here, I will highlight the work performed in my research group on using these approaches to accelerate natural product discovery, as well as to study microbe-microbe and host-microbe interactions in human, plant and animal microbiomes. Specifically, I will discuss the use of computational approaches to investigate biosynthetic diversity across large numbers of genomes, and integrative genome/transcriptome/metabolome mining to associate gene clusters to molecules and ecological functions in order to elucidate the molecular basis of microbiome-associated phenotypes.

Urinary tract infections (UTIs) caused by uropathogenic Escherichia coli (UPEC) belong to the most common bacterial infections in humans. A crucial infection step is the adhesion of type 1-piliated UPEC to mannosylated glycoproteins on uroepithelial cells. Uromodulin (UMOD) is the most abundant protein in human urine and forms long homopolymeric filaments that antagonize UPEC adhesion, however, little was known about its structure and protective mechanism.

By using an integrative approach of glyco-mass spectrometry, light microscopy, cryo-focused ion beam milling and cryo-electron tomography (cryoET), we resolved the architecture of UMOD filaments and could show how UMOD serves as a multivalent antagonist against UPEC adhesion directly in patient samples. CryoET and subtomogram averaging revealed that UMOD filaments are composed of a zigzag-shaped core module formed by the zona pellucida (ZP) domains and laterally protruding arms consisting of the N-terminal UMOD segment. We further demonstrate that UMOD forms a loose network of filaments around individual piliated bacteria and mediates the aggregation of bacteria by a dense mesh. These higher order structures likely prevent UPEC adhesion to the urinary epithelium and favor efficient clearance by micturition, representing a previously unrecognized aspect of UMOD biology. Analyzing fresh urine from patients with an acute UTI by light microscopy and cryoET confirmed our in vitro results and corroborated UMOD’s universal protective mechanism against pathogens. This demonstrates the power of cryoET to assess the molecular basis of a disease directly in patient samples.

Reference

Weiss, G.L., Stanisich, J.J., Sauer, M.M., Lin, C.-W., Eras, J., Zyla, D.S., Trück, J., Devuyst, O., Aebi, M., Pilhofer, M., et al. (2020). Architecture and function of human uromodulin filaments in urinary tract infections. Science 369, 1005–1010.

The largest organ of the human body is skeletal muscle, accounting for up to 50% of the entire body mass. Muscle mass and function is tightly controlled and can be lost as a consequence of pathological damage to the neuromuscular system, immobility or aging. My laboratory has studied mechanisms involved in the development and maintenance of the neuromuscular system over several decades. In particular, we are interested in how the synapse between motor neurons and muscle fibers is formed and maintained. In one of our projects, we hypothesized that extracellular matrix proteins with essential functions at the nerve-muscle synapse could be engineered to replace the function of laminin-α2, the loss of which causes a rare, early-onset muscular dystrophy. Using transgenic mouse models of the disease, we indeed demonstrated that expression of the engineered proteins strongly ameliorated the muscular dystrophy. Most strikingly, lifespan of treated mice reached more than two years whereas non-treated mice usually die within the first 4 months of life. These experiments serve as the basis for our current efforts to translate this approach into clinics using adeno-associated virus-mediated gene therapy. 

The centriole (also called basal body) is an evolutionary conserved organelle that is typically 500 nm long and 250 nm in diameter and composed of about 100 different proteins, all in multiple copies. How these proteins are organized at the level of the centriolar architecture is poorly understood and only a fraction of these proteins have been clearly studied and precisely localized. Moreover, a clear molecular overview of the centriolar structure and assembly is currently lacking. I will present the latest work from my laboratory that uses several approaches such as in vitro reconstruction, in situ cryo-tomography and high resolution molecular mapping using U-ExM to address these fundamental questions.

The novel coronavirus SARS-CoV-2, the causative agent of COVID-19 respiratory disease is evolving during the current pandemic.  New variants show enhanced replication and the potential to evade therapeutic antibodies.  In the near future, variants may even evade first generation vaccines.  The currently approved direct acting antiviral remdesivir targets the viral RNA-dependent RNA polymerase which is subject to rapid evolution as it is encoded by the viral RNA genome.  In order to develop therapeutic approaches which act in a pan-coronavirus manner we and our colleagues at the QBI Coronavirus Research Group (QCRG) have mapped the human proteins (host factors) which multiple Coronaviruses rely on for replication.  Through a rapid drug repurposing effort we have identified zotatifin, a clinical eIF4A inhibitor as a host factor targeted therapeutic.  Zotatifin which is based on the natural product rocaglamide A works as a molecular glue to trap eIF4A on its target, the (+) RNA viral genome.  Other examples of targeting essential host factors, including those for immune evasion will be discussed.