Mutations in the receptor-binding domain of human SARS CoV-2 spike protein increases its affinity to bind human ACE-2 receptor

The severe acute respiratory syndrome virus-2 (SARS CoV-2) infection has resulted in the current global pandemic. The binding of SARS CoV-2 spike protein receptor-binding domain (RBD) to the human angiotensin converting enzyme-2 (ACE-2) receptor causes the host infection. The spike protein has undergone several mutations with reference to the initial strain isolated during December 2019 from Wuhan, China. A number of these mutant strains have been reported as variants of concern and as variants being monitored. Some of these mutants are known to be responsible for increased transmissibility of the virus.
The reason for the increased transmissibility caused by the point mutations can be understood by studying the structural implications and inter-molecular interactions in the binding of viral spike protein RBD and human ACE-2. Here, we use the crystal structure of the RBD in complex with ACE-2 available in the public domain and analyse the 250 ns molecular dynamics (MD) simulations of wild-type and mutants; K417N, K417T, N440K, N501Y, L452R, T478K, E484K and S494P.
The ionic, hydrophobic and hydrogen bond interactions, amino acid residue flexibility, binding energies and structural variations are characterized. The MD simulations provide clues to the molecular mechanisms of ACE-2 receptor binding in wild-type and mutant complexes. The mutant spike proteins RBD were associated with greater binding affinity with ACE-2 receptor. Communicated by Ramaswamy H. Sarma.

COVID-19 and Alzheimer’s disease: Meninges-mediated neuropathology

SARS-CoV-2 the causative agent of COVID-19 displays a broad range of pathophysiology. Cytokine storms associated with COVID-19 damage the blood-brain barrier (BBB) and allow pro-inflammatory factors to invade the brain, further promoting neurodegeneration. While SARS-CoV-2 viral RNA and proteins have been detected in brain tissues, the mechanisms of neuroinvasion remain unknown. COVID-19 has had a disproportionate impact on those suffering from neurodegenerative disorders such as Alzheimer’s disease (AD).
Understanding the mechanisms of SARS-CoV-2 neuroinvasion is crucial to study the long-term neurocognitive effects of COVID-19 on AD pathology.
Viruses can infiltrate the brain through the meninges via infected immune cells. The meninges regulate the immune surveillance of the brain and play a key role in the efflux of pathogens from the brain. Meningeal dysfunction has been demonstrated to exacerbate amyloid-beta pathogenesis. In this study, we explore the neuroinvasion pathway of SARS-CoV-2 through the meninges and its effect on AD pathology.
Method: 5x FAD x hACE2 mice were inoculated intranasally with a sublethal dose of SARS-CoV-2. The mice were maintained for 2 weeks. Mouse brains and meninges were harvested. The tissue was stained and immunofluorescence imaging was conducted to study viral proliferation and immune responses. Histo-cytometry was conducted for quantitative imaging analysis. Gene expression studies were done using Nanostring assays. All experiments involving the SARS-Cov-2 virus were carried out in a BSL3 facility.
Result: This ongoing study demonstrates the proliferation of the SARS-CoV-2 virus in the brain via meningeal lymphatics. SARS-CoV-2 infection resulted in increased neuroinflammation. Additionally, inflammatory responses induced meningeal dysfunction resulting in increased amyloid-beta pathology and cerebrospinal fluid drainage.
Conclusion: Given the increasing evidence for a viral hypothesis of Alzheimer’s Disease it is extremely important to study the neurodegenerative effects of COVID-19 which has affected millions worldwide. We demonstrate that SARS-CoV-2 infiltrates the brain via the meninges promoting neuroinflammation. Furthermore, amyloid-beta pathologies are exacerbated by COVID-19 in animal models providing preclinical evidence of the long-term neurodegenerative effects of COVID-19.

Exosomes Recovered From the Plasma of COVID-19 Patients Expose SARS-CoV-2 Spike-Derived Fragments and Contribute to the Adaptive Immune Response

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by beta-coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that has rapidly spread across the globe starting from February 2020. It is well established that during viral infection, extracellular vesicles become delivery/presenting vectors of viral material. However, studies regarding extracellular vesicle function in COVID-19 pathology are still scanty. Here, we performed a comparative study on exosomes recovered from the plasma of either MILD or SEVERE COVID-19 patients.
We show that although both types of vesicles efficiently display SARS-CoV-2 spike-derived peptides and carry immunomodulatory molecules, only those of MILD patients are capable of efficiently regulating antigen-specific CD4+ T-cell responses. Accordingly, by mass spectrometry, we show that the proteome of exosomes of MILD patients correlates with a proper functioning of the immune system, while that of SEVERE patients is associated with increased and chronic inflammation.
Overall, we show that exosomes recovered from the plasma of COVID-19 patients possess SARS-CoV-2-derived protein material, have an active role in enhancing the immune response, and possess a cargo that reflects the pathological state of patients in the acute phase of the disease.

Role of SARS-CoV-2 in causing blood-brain barrier leakage and microglial activation as a risk factor of cognitive deterioration in subjects at risk of Alzheimer’s disease

The recent pandemic provides evidence of altered central nervous system (CNS) function in response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). SARS-COV-2 invades the CNS by binding angiotensin-converting enzyme 2 (ACE2) expressed on neurons and glia. SARS-COV-2 may have effects on increased permeability of endothelial cells within the blood-brain barrier (BBB) as studies have shown that the S1 protein can transverse the BBB.
This is interesting because leakage of the BBB is implicated in Alzheimer’s disease (AD) pathogenesis. We hypothesized that SARS-CoV-2 infection leads to innate stimulated inflammation, ultimately activating microglial cells and an influx of pro-inflammatory cytokines and leukocytes in the meninges, contributing to increased permeability of the BBB. This BBB permeability increases AD susceptibility in subjects at risk by causing irreversible damage to the BBB and microglial cell activation.
Method: We developed a double transgenic mouse model using mice expressing human ACE2 receptor and 5xFAD mice that exhibit increased neuropathology seen in human AD allowing modeling of AD in SARS-CoV-2 pathogenesis. The hACE2/5xFAD double transgenic mice were intranasally inoculated with a sub-lethal dose of SARS-CoV-2 to test the hypothesis that SARS-COV-2 potentiates AD pathology and cognitive deterioration through impairment of the BBB. Leukocyte and cytokine populations were measured by flow cytometry and single-nuclei RNA sequencing of the meninges for characterization of microglial populations.
Result: SARS-CoV-2 creates a cytotoxic environment in the brain immediately following infection in hACE2/5xFAD mice leading to leakage of the BBB in the meninges. Activation of microglial innate cells by SARS-COV-2 invasion of the CNS will cause neural deterioration having long term implications on cognitive function. The hACE2/5xFAD mouse model allows us to uncover implications for SARS-COV-2 on AD cognitive deterioration.
Conclusion: The hACE2/5xFAD mouse allows modeling of SARS-CoV-2 in developing AD cases and allowed us to determine the immune environment generated in the meninges in response to SARS-CoV-2 infections. This mouse model provides a platform to proactively determine the effects of SARS-CoV-2 in developing AD cases, a methodology to be exploited for future mouse models determining the relationship of other viruses on AD pathology, and the opportunity to address phenotypes with therapeutics for preventative initiatives.

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