Age-dependent appearance of SARS-CoV-2 entry sites in mouse chemosensory systems reflects COVID-19 anosmia-ageusia symptoms

AbstractCOVID-19 pandemic has given rise to a collective scientific effort to study its viral causing agent SARS-CoV-2. Research is focusing in particular on its infection mechanisms and on the associated-disease symptoms. Interestingly, this environmental pathogen directly affects the human chemosensory systems leading to anosmia and ageusia. Evidence for the presence of the cellular entry sites of the virus, the ACE2/TMPRSS2 proteins, has been reported in non-chemosensory cells in the rodent’s nose and mouth, missing a direct correlation between the symptoms reported in patients and the observed direct viral infection in human sensory cells. Here, mapping the gene and protein expression of ACE2/TMPRSS2 in the mouse olfactory and gustatory cells, we precisely identify the virus target cells to be of basal and sensory origin and reveal the age-dependent appearance of viral entry-sites. Our results propose an alternative interpretation of the human viral-induced sensory symptoms and give investigative perspectives on animal models.

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Age-dependent appearance of SARS-CoV-2 entry cells in mouse chemosensory systems reflects COVID-19 anosmia and ageusia symptoms.

10.1101/2021.03.29.437530 ◽

2021 ◽

Author(s):

Julien Brechbuhl ◽

Dean Wood ◽

Sofiane Bouteiller ◽

Ana Catarina Lopes ◽

Chantal Verdumo ◽

...

Keyword(s):

Mouse Models ◽

Sensory Cells ◽

Target Cells ◽

Humanized Mouse ◽

Disease Symptoms ◽

Gene And Protein Expression ◽

Age Dependent ◽

Infection Mechanisms ◽

Humanized Mouse Models ◽

Chemosensory Systems

COVID-19 pandemic has given rise to a collective scientific effort to study its viral causing agent SARS-CoV-2. Research is focusing in particular on its infection mechanisms and on the associated-disease symptoms. Interestingly, this environmental pathogen directly affects the human chemosensory systems leading to anosmia and ageusia. Evidence for the presence of the cellular entry sites of the virus, the ACE2 and the TMPRSS2 proteins, has been reported in non-chemosensory cells in the nose and mouth of the rodents, missing a direct correlation between the symptoms reported in patients and the observed direct viral infection in human sensory cells. Here, mapping the gene and protein expression of ACE2 and TMPRSS2 in the mouse olfactory and gustatory cells, we precisely identified the virus target cells to be of basal and sensory origin and we revealed their age-dependent appearance. Our results not only clarify human viral-induced sensory symptoms but also propose new investigative perspectives based on ACE2-humanized mouse models.

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Repurposing Nimesulide, a Potent Inhibitor of the B0AT1 Subunit of the SARS-CoV-2 Receptor, as a Therapeutic Adjuvant of COVID-19

SLAS DISCOVERY Advancing Life Sciences ◽

10.1177/2472555220934421 ◽

2020 ◽

Vol 25 (10) ◽

pp. 1171-1173 ◽

Cited By ~ 4

Author(s):

Mariafrancesca Scalise ◽

Cesare Indiveri

Keyword(s):

Viral Entry ◽

3D Structure ◽

World Health ◽

Effective Vaccine ◽

Target Cells ◽

Physical Interaction ◽

Disease Symptoms ◽

Angiotensin Converting Enzyme 2 ◽

Global Pandemic ◽

Health Organization

The global pandemic caused by the SARS-CoV-2 infection is a health emergency that needs to be addressed immediately. The international scientific community, following World Health Organization (WHO) indications, launched different trials for testing drugs putatively able to block the SARS-CoV-2 infection or treat the COVID-19 disease symptoms. In parallel, studies devoted to a better understanding of SARS-CoV-2 biology are in the course for designing an effective vaccine. One of the human membrane proteins known to be docked by the virus is angiotensin-converting enzyme 2 (ACE2), proposed to be responsible for viral entry in target cells. Recently, the 3D structure of ACE2 has been obtained, showing its physical interaction with B0AT1 (SLC6A19), a plasma membrane transporter involved in the trafficking of amino acids in cells. The receptor targeted by SARS-CoV-2 is a supercomplex formed by a dimer of ACE2-B0AT1, in which ACE2 binds the viral protein and B0AT1 stabilizes the heterodimer. As a serendipity occurrence, nimesulide was shown to abolish the transport function of B0AT1. Here we suggest including nimesulide in the list of drugs to be tested for the identification of co-adjuvants in the treatment of COVID-19.

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SARS-CoV-2 Cellular Infection and Therapeutic Opportunities: Lessons Learned from Ebola Virus

Membranes ◽

10.3390/membranes11010064 ◽

2021 ◽

Vol 11 (1) ◽

pp. 64

Author(s):

Jordana Muñoz-Basagoiti ◽

Daniel Perez-Zsolt ◽

Jorge Carrillo ◽

Julià Blanco ◽

Bonaventura Clotet ◽

...

Keyword(s):

Viral Entry ◽

Ebola Virus ◽

Lessons Learned ◽

Productive Infection ◽

Target Cells ◽

Emerging Viruses ◽

Highly Pathogenic ◽

Life Threatening ◽

Pathogenic Viruses ◽

Cellular Machinery

Viruses rely on the cellular machinery to replicate and propagate within newly infected individuals. Thus, viral entry into the host cell sets up the stage for productive infection and disease progression. Different viruses exploit distinct cellular receptors for viral entry; however, numerous viral internalization mechanisms are shared by very diverse viral families. Such is the case of Ebola virus (EBOV), which belongs to the filoviridae family, and the recently emerged coronavirus SARS-CoV-2. These two highly pathogenic viruses can exploit very similar endocytic routes to productively infect target cells. This convergence has sped up the experimental assessment of clinical therapies against SARS-CoV-2 previously found to be effective for EBOV, and facilitated their expedited clinical testing. Here we review how the viral entry processes and subsequent replication and egress strategies of EBOV and SARS-CoV-2 can overlap, and how our previous knowledge on antivirals, antibodies, and vaccines against EBOV has boosted the search for effective countermeasures against the new coronavirus. As preparedness is key to contain forthcoming pandemics, lessons learned over the years by combating life-threatening viruses should help us to quickly deploy effective tools against novel emerging viruses.

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Prevention of Measles Virus Infection by Intranasal Delivery of Fusion Inhibitor Peptides

Journal of Virology ◽

10.1128/jvi.02417-14 ◽

2014 ◽

Vol 89 (2) ◽

pp. 1143-1155 ◽

Cited By ~ 28

Author(s):

C. Mathieu ◽

D. Huey ◽

E. Jurgens ◽

J. C. Welsch ◽

I. DeVito ◽

...

Keyword(s):

Animal Models ◽

Measles Virus ◽

Structural Transition ◽

Specific Treatment ◽

Heptad Repeat ◽

Receptor Interaction ◽

Target Cells ◽

Inhibitory Peptides ◽

Fusion Inhibitors

ABSTRACTMeasles virus (MV) infection is undergoing resurgence and remains one of the leading causes of death among young children worldwide despite the availability of an effective measles vaccine. MV infects its target cells by coordinated action of the MV H and the fusion (F) envelope glycoprotein; upon receptor engagement by H, the prefusion F undergoes a structural transition, extending and inserting into the target cell membrane and then refolding into a postfusion structure that fuses the viral and cell membranes. By interfering with this structural transition of F, peptides derived from the heptad-repeat (HR) regions of F can potently inhibit MV infection at the entry stage. We show here that specific features of H's interaction with its receptors modulate the susceptibility of MV F to peptide fusion inhibitors. A higher concentration of inhibitory peptides is required to inhibit F-mediated fusion when H is engaged to its nectin-4 receptor than when H is engaged to its CD150 receptor. Peptide inhibition of F may be subverted by continued engagement of receptor by H, a finding that highlights the ongoing role of H-receptor interaction after F has been activated and that helps guide the design of more potent inhibitory peptides. Intranasal administration of these peptides results in peptide accumulation in the airway epithelium with minimal systemic levels of peptide and efficiently prevents MV infectionin vivoin animal models. The results suggest an antiviral strategy for prophylaxis in vulnerable and/or immunocompromised hosts.IMPORTANCEMeasles virus (MV) infection causes an acute illness that may be associated with infection of the central nervous system (CNS) and severe neurological disease. No specific treatment is available. We have shown that parenterally delivered fusion-inhibitory peptides protect mice from lethal CNS MV disease. Here we show, using established small-animal models of MV infection, that fusion-inhibitory peptides delivered intranasally provide effective prophylaxis against MV infection. Since the fusion inhibitors are stable at room temperature, this intranasal strategy is feasible even outside health care settings, could be used to protect individuals and communities in case of MV outbreaks, and could complement global efforts to control measles.

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Age-Dependent Consequences of Status Epilepticus: Animal Models

Epilepsia ◽

10.1111/j.1528-1167.2007.01069.x ◽

2007 ◽

Vol 48 (s2) ◽

pp. 75-82 ◽

Cited By ~ 38

Author(s):

Morris H. Scantlebury ◽

James G. Heida ◽

Henry J. Hasson ◽

Jana Velíšková ◽

Libor Velíšek ◽

...

Keyword(s):

Status Epilepticus ◽

Animal Models ◽

Age Dependent

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Induction of long-term facilitation in Aplysia sensory neurons by local application of serotonin to remote synapses

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.90.23.11411 ◽

1993 ◽

Vol 90 (23) ◽

pp. 11411-11415 ◽

Cited By ~ 35

Author(s):

G A Clark ◽

E R Kandel

Keyword(s):

Gene Expression ◽

Sensory Neurons ◽

Cell Body ◽

Sensory Cells ◽

Target Cells ◽

Synaptic Terminals ◽

Dissociated Cell Culture ◽

Synaptic Changes ◽

Long Term Facilitation

Long-term synaptic facilitation at the connections of Aplysia sensory neurons onto their target cells involves alterations in gene expression. How then are the relevant cellular signals for the induction and expression of long-term synaptic changes conveyed between the nucleus and remote synaptic terminals? We have explored this question using a set of remote, peripheral terminals of siphon sensory cells, which are approximately 3 cm from the sensory cell body in the abdominal ganglion. We found that these remote synapses, like the proximal synapses previously studied in dissociated cell culture, can exhibit long-term facilitation 24 hr after cell-wide serotonin application. Furthermore, serotonin applications restricted to the remote synaptic terminals nevertheless produced long-term facilitation, indicating that signals generated in synaptic regions can trigger the long-term process, perhaps via retrograde signals to the nucleus to modify gene expression, followed by anterograde signals back to the terminal. Serotonin applications restricted to the cell body and proximal synapses of the sensory neuron also produced long-term facilitation at remote synapses, although to a lesser extent, suggesting that long-term facilitation is expressed cell-wide, but that superimposed on this cell-wide facilitation there appears to be a component that is synapse-specific.

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Middle East Respiratory Syndrome Coronavirus Spike Protein Is Not Activated Directly by Cellular Furin during Viral Entry into Target Cells

Journal of Virology ◽

10.1128/jvi.00683-18 ◽

2018 ◽

Vol 92 (19) ◽

Cited By ~ 30

Author(s):

Shutoku Matsuyama ◽

Kazuya Shirato ◽

Miyuki Kawase ◽

Yutaka Terada ◽

Kengo Kawachi ◽

...

Keyword(s):

Middle East ◽

Viral Entry ◽

Cathepsin L ◽

Inhibitory Effect ◽

Middle East Respiratory Syndrome ◽

Cell Entry ◽

Target Cells ◽

Furin Cleavage ◽

S Protein ◽

Entry Assay

ABSTRACT Middle East respiratory syndrome coronavirus (MERS-CoV) utilizes host cellular proteases to enter cells. A previous report shows that furin, which is distributed mainly in the Golgi apparatus and cycled to the cell surface and endosomes, proteolytically activates the MERS-CoV spike (S) protein following receptor binding to mediate fusion between the viral and cellular membranes. In this study, we reexamined furin usage by MERS-CoV using a real-time PCR-based virus cell entry assay after inhibition of cellular proteases. We found that the furin inhibitor dec-RVKR-CMK blocked entry of MERS-CoV harboring an S protein lacking furin cleavage sites; it even blocked entry into furin-deficient LoVo cells. In addition, dec-RVKR-CMK inhibited not only the enzymatic activity of furin but also those of cathepsin L, cathepsin B, trypsin, papain, and TMPRSS2. Furthermore, a virus cell entry assay and a cell-cell fusion assay provided no evidence that the S protein was activated by exogenous furin. Therefore, we conclude that furin does not play a role in entry of MERS-CoV into cells and that the inhibitory effect of dec-RVKR-CMK is specific for TMPRSS2 and cathepsin L rather than furin. IMPORTANCE Previous studies using the furin inhibitor dec-RVKR-CMK suggest that MERS-CoV utilizes a cellular protease, furin, to activate viral glycoproteins during cell entry. However, we found that dec-RVKR-CMK inhibits not only furin but also other proteases. Furthermore, we found no evidence that MERS-CoV uses furin. These findings suggest that previous studies in the virology field based on dec-RVKR-CMK should be reexamined carefully. Here we describe appropriate experiments that can be used to assess the effect of protease inhibitors on virus cell entry.

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Structure, function and antigenicity of the SARS-CoV-2 spike glycoprotein

10.1101/2020.02.19.956581 ◽

2020 ◽

Cited By ~ 91

Author(s):

Alexandra C. Walls ◽

Young-Jun Park ◽

M. Alexandra Tortorici ◽

Abigail Wall ◽

Andrew T. McGuire ◽

...

Keyword(s):

Receptor Binding ◽

Viral Entry ◽

Neutralizing Antibodies ◽

Target Cells ◽

Furin Cleavage ◽

Humoral Immune ◽

Binding Domains ◽

Furin Cleavage Site ◽

Recent Emergence ◽

Novel Coronavirus

SUMMARYThe recent emergence of a novel coronavirus associated with an ongoing outbreak of pneumonia (Covid-2019) resulted in infections of more than 72,000 people and claimed over 1,800 lives. Coronavirus spike (S) glycoprotein trimers promote entry into cells and are the main target of the humoral immune response. We show here that SARS-CoV-2 S mediates entry in VeroE6 cells and in BHK cells transiently transfected with human ACE2, establishing ACE2 as a functional receptor for this novel coronavirus. We further demonstrate that the receptor-binding domains of SARS-CoV-2 S and SARS-CoV S bind with similar affinities to human ACE2, which correlates with the efficient spread of SARS-CoV-2 among humans. We found that the SARS-CoV-2 S glycoprotein harbors a furin cleavage site at the boundary between the S1/S2 subunits, which is processed during biogenesis and sets this virus apart from SARS-CoV and other SARS-related CoVs. We determined a cryo-electron microscopy structure of the SARS-CoV-2 S ectodomain trimer, demonstrating spontaneous opening of the receptor-binding domain, and providing a blueprint for the design of vaccines and inhibitors of viral entry. Finally, we demonstrate that SARS-CoV S murine polyclonal sera potently inhibited SARS-CoV-2 S-mediated entry into target cells, thereby indicating that cross-neutralizing antibodies targeting conserved S epitopes can be elicited upon vaccination.

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Molecular Signaling and Cellular Pathways for Virus Entry

ISRN Virology ◽

10.5402/2013/306595 ◽

2013 ◽

Vol 2013 ◽

pp. 1-8 ◽

Cited By ~ 1

Author(s):

Pei-I Chi ◽

Hung-Jen Liu

Keyword(s):

Signaling Pathways ◽

Viral Entry ◽

Virus Entry ◽

Host Cells ◽

Target Cells ◽

Molecular Signaling ◽

Cellular Processes ◽

Cellular Pathways ◽

Host Life ◽

Endocytic Vesicles

The cell signaling plays a pivotal role in regulating cellular processes and is often manipulated by viruses as they rely on the functions offered by cells for their propagation. The first stage of their host life is to pass the genetic materials into the cell. Although some viruses can directly penetrate into cytosol, in fact, most virus entry into their host cells is through endocytosis. This machinery initiates with cell type specific cellular signaling pathways, and the signaling compounds can be proteins, lipids, and carbohydrates. The activation can be triggered in a very short time after virus binds on target cells, such as receptors. The signaling pathways involved in regulation of viral entry are wide diversity that often cross-talk between different endocytosis results. Furthermore, some viruses have the ability to use the multiple internalization pathways which leads to the regulation being even more complex. In this paper, we discuss some recent advances in our understanding of cellular pathways for virus entry, molecular signaling during virus entry, formation of endocytic vesicles, and the traffic.

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OR28-01 Constitutive Activation of NRF2 Antioxidant Response Leads to Age-Dependent Goiter and Compensated Hypothyroidism in Male Mice

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa046.2042 ◽

2020 ◽

Vol 4 (Supplement_1) ◽

Author(s):

Dionysios Chartoumpekis ◽

Panos Ziros ◽

Cédric Renaud ◽

Massimo Bongiovanni ◽

Ioannis Habeos ◽

...

Keyword(s):

Thyroid Function ◽

Expression Profiles ◽

Adult Life ◽

Model Organisms ◽

Mrna Levels ◽

Loss Of Function ◽

Gene And Protein Expression ◽

Age Dependent ◽

Diffuse Goiter ◽

Early Adult Life

Abstract Background: Familial non-toxic multinodular goiter (MNG) is a rare disease. KEAP1 gene (Kelch-like ECH-associated protein 1) that encodes the main inhibitor of nuclear factor erythroid 2-related transcription factor 2 (Nrf2), a central mediator of antioxidant responses, has been found to be one of the mutated genes that lead to familial MNG. The proposed association of KEAP1 with familial MNG is based on only two loss-of-function mutations in respective Japanese families, only one of which included proper phenotyping and demonstration of co-segregation of phenotype and mutation. To date, there is no experimental evidence from model organisms to support that decreased Keap1 levels can cause goiter. Hypothesis: We hypothesized that enhanced Nrf2 signaling induced by loss of Keap1 function in mice can lead to goiter. Methods: To this end, male Keap1 hypomorphic C57BL/6J mice that express ~80% less Keap1 in their tissues (Keap1 knockdown mice:“Keap1KD”) were studied at 3 and 12 months of age and compared to wild-type mice (WT). Plasma, thyroids and pituitary glands were collected for assessment of thyroid function by radioimmunoassays and for histology as well as gene and protein expression by quantitative PCR and immunoblotting respectively. Results: Keap1KD showed diffuse goiter that began to develop in early adult life and became highly prominent at the age of 12 months when the thyroids of Keap1KD were 6-fold heavier than WT. Histomorphometry assessment of thyroids showed that Keap1KD had ~3-fold larger follicle area and colloid compartment but no thyroid nodules or hyperplasia was detected. Keap1KD also showed primary hypothyroidism already in early adult life that was eventually well-compensated over time by increased TSH levels (at age of 12 months: WT TSH=47.7±9.1 mU/L, Keap1KD TSH=460±74 mU/L). This was also reflected in the pituitary gland of Keap1KD where Tshb mRNA was ~3-fold higher than WT. Despite a known stimulatory effect of Nrf2 on Tg gene transcription and Tg protein abundance, these measures were decreased in the thyroid of Keap1KD mice. No clear patterns were observed in the expression profiles of other thyroid hormone synthesis-specific factors, such as Duox1, Duoxa1, Duox2, Duoxa2, Tpo, Nis, Dio1, Dio2, Dehal1 mRNA levels, with the exception of Tg-processing and Tg-degrading cathepsins, including an increase in mature forms of cathepsins D, L and S. Conclusions: Keap1KD mice showed age-dependent diffuse goiter and compensated hypothyroidism. The precise mechanism accounting for the thyroidal phenotype remains to be elucidated, but it may involve enhanced Tg solubilization and excessive lysosomal Tg degradation. This study unravels novel roles of the druggable Keap1/Nrf2 pathway in thyroid function and economy. Subclinical hypothyroidism in Keap1KD mice may have broader implications regarding their use in metabolic research.

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