Shaker, M. | Search bioRxiv

bioRxiv2026

Towards Understanding the Drivers of Antibody-Antigen Binding

Antibody therapeutics are capable of binding to target antigens with a high degree of specificity and affinity. Gaining an understanding of how antibody-antigen interactions are governed can provide valuable insights that may assist with rational paratope design and epitope prediction. In this work, we apply the FTMap algorithm to systematically characterize binding hot spots - regions on a protein surface that contribute disproportionately to molecular recognition, to a set of 50 antibody-antigen complexes. From our analysis, we find that interface hot spots are typically concentrated on the paratope (antibody side) of the interface, indicating that paratopes typically function as hot spot rich environments in which the antigen can bind. Additionally, we observe that hot spot formation on both sides of the interface is particularly enriched by Trp and Tyr residues, underscoring the key role of aromatic side chains with some amphiphilic character in antibody design. Furthermore, we find that when strong interface hot spots are detected, they tend to persist in the apo conformation, suggesting that there is an inherent structural stability that surrounds core interface hot spots. These findings demonstrate the utility of computational solvent mapping for analyzing protein-protein interfaces, and highlights that at least in most cases antibodies drive antibody-antigen interactions. Statement of SignificanceAntibodies represent an important and expanding class of therapeutics for a range of diseases including infectious diseases, cancers, and autoimmune disorders. Enhancing our fundamental understanding of what drives antibody-antigen interactions is critical to our enhancing our ability to modulate these interactions. This work presents a systematic, physics-based study of antibody-antigen interfaces, and identifies key drivers of binding.

bioinformatics↗

bioRxiv2024

SARS-CoV-2 infection in hiPSC-derived neurons is cathepsin-dependent and causes accumulation of HIF1alpha and phosphorylated tau

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been shown to infect the human brain and a subset of human neurons in vitro. We have previously demonstrated that the virus enters the human induced pluripotent stem cell (hiPSC)-derived neurons via an endosomal-lysosomal pathway, which is dependent on low levels of angiotensin-converting enzyme 2 (ACE2) and independent of transmembrane serine protease 2 (TMPRSS2). Here, we use hiPSC-derived neurons overexpressing ACE2 in co-culture with human astrocytes to show that the infection with both SARS-CoV-2 Wuhan and Omicron XBB.1.5 variants is dependent on cathepsins and can be efficiently blocked by an inhibitor of cathepsin B (CA-074-ME). The result was reproducible in non-transgenic hiPSC-derived cortical organoids. The cathepsin L inhibitor SB412515 was less effective against the Wuhan strain but equally effective against the Omicron variant. Using PCR and reinfection assays, we show that SARS-CoV-2 can replicate in neurons in 2D co-cultures. Interestingly, the infectivity of the newly produced virions declined at 24 hours post-infection despite a further increase in released viral RNA at later time points, suggesting the possible activation of an antiviral response in neurons and/or astrocytes, which is supported by a correspondent increase in the levels of secreted cytokines. Furthermore, the number of infected neurons decreased within five days, suggesting that SARS-CoV-2 infection eventually leads to the death of the target neuronal cell in vitro. The infection also caused the accumulation of the hypoxia-inducible stress factor HIF1- in infected neurons under normoxia. Finally, we confirm and expand the previous finding that in SARS-CoV-2 infected neurons, the microtubule-associated protein tau is hyperphosphorylated at multiple loci, including S202/T205, and mislocalized to the soma of the infected neurons. Hyperphosphorylation and mislocalization of tau are hallmarks of Alzheimers disease (AD) and other tauopathies. Our data provides further evidence supporting the neurodegenerative potential of SARS-CoV-2 infection. SummaryThe recent COVID-19 pandemic has raised concerns about the potential for SARS-CoV-2 to infect the brain and worsen brain diseases like Alzheimers disease. Research has shown that SARS-CoV-2 can indeed infect the human brain, including a small number of neurons and other brain cells in laboratory settings. In our previous studies, we identified the endosomal pathway as the route the virus uses to enter neuronal cells. In this study, we build on that work by demonstrating that inhibitors of endo-lysosomal cathepsin proteases can block this neuronal infection. We also found that infectious progeny virions are released from the infected neuronal cells. Importantly, the infection proves harmful to the host cells, as evidenced by a decrease in the number of infected cells in experimental cultures over a five-day period. Additionally, we confirm and expand on earlier findings that SARS-CoV-2 infection leads to the phosphorylation and altered localization of the tau protein, a process associated with brain diseases like Alzheimers. Finally, we observed an increase in the production of inflammatory cytokines following neuronal infection with SARS-CoV-2, along with an accumulation of the stress marker protein HIF-1 in neurons. This protein has been linked to other viral infections and Alzheimers disease. Overall, our data suggest that SARS-CoV-2 exhibits neurodegenerative characteristics.

microbiology↗

bioRxiv2023

Rapamycin mitigates Valproic Acid-induced teratogenicity in human and animal models by suppressing AP-1-mediated senescence

Valproic acid (VPA) is an effective and widely used anti-seizure medication but is teratogenic when used during pregnancy, affecting brain and spinal cord development for reasons that remain largely unclear. Here we designed a genetic recombinase-based SOX10 reporter system in human pluripotent stem cells that enables tracking and lineage tracing of Neural Crest cells (NCCs) in a human organoid model of the developing neural tube. We found that VPA induces extensive cellular senescence and promotes mesenchymal differentiation of human NCCs at the expense of neural lineages. We next show that the clinically-approved drug, Rapamycin, inhibits AP1-mediated senescence and restores aberrant NCC differentiation trajectory in human organoids exposed to VPA. Notably, in vivo validation in developing zebrafish highlighted the therapeutic promise of this approach. Collectively our data identifies a novel mechanism for VPA-associated neurodevelopmental teratogenicity and a potential pharmacological preventative strategy. The results exemplify the power of genetically modified human stem cell-derived organoid models for drug discovery and safety testing.

neuroscience↗