Biology subjects

Bouma, E. M.

Publications and source records attributed to Bouma, E. M..

6 recordsLinked to original sources

bioRxiv2026

The Heat shock protein 70 machinery is crucial in the production of infectious chikungunya virus progeny

Over the past decades, chikungunya virus (CHIKV), a re-emerging arthropod-borne alphavirus, has caused outbreaks in many (sub)tropical regions, but also in more temperate regions of the world, including Europe. CHIKV poses a significant health burden due to high infection rates during epidemics, symptom progression into chronic arthritic manifestations, and the lack of specific antiviral treatments. Multiple studies have shown that several viruses rely on host molecular chaperones, particularly the central Heat shock protein 70 (Hsp70), for replication. Hsp70s are guided by co-chaperones that drive their functionality and specificity. Here, we used chemical inhibitors of Hsp70-co-chaperone interactions to study the role of this molecular chaperone machinery in CHIKV replication. Our findings revealed that Hsp70 inhibition significantly reduces the CHIKV infectious particle production without affecting host-cell viability. Inhibition of the Hsp70-co-chaperone interaction primarily impedes the post-RNA replication stages of the CHIKV infectious cycle, affecting viral protein expression and reducing both the number and infectivity of released virions. Moreover, Hsp70 inhibition displayed antiviral activity in skin explants. Together, these results suggest that targeting the Hsp70 network could be a viable antiviral strategy against CHIKV infections. Author summaryAs obligatory intracellular parasites, viruses rely entirely on the machinery of the host cell to produce new viral particles. In our study, we investigated whether a specific group of host proteins, known as molecular chaperones, is important for the replication of chikungunya virus (CHIKV), a reemerging mosquito-borne virus that can cause long-lasting joint pains and lacks specific antiviral treatments. We focused on one key family of molecular chaperones, called Heat shock protein 70s (Hsp70s), which supports protein folding and quality control in cells. We used chemical compounds to block Hsp70 function and observed that CHIKV replication was strongly reduced, while host cells remained healthy. We found that Hsp70 is especially important in the later stages of the virus life cycle, where it helps produce viral proteins and new infectious virus particles. When Hsp70 was blocked, fewer and less infectious virus particles were produced. We also showed that this effect holds true in biopsies of mouse skin tissue, which mimics the initial site of infection. These findings identify Hsp70 as an important host factor for CHIKV that may serve as a potential target for new antiviral therapies.

microbiology↗

bioRxiv2021

Moxidectin and ivermectin inhibit SARS-CoV-2 replication in Vero E6 cells but not in human primary airway epithelium cells

Antiviral therapies are urgently needed to treat and limit the development of severe COVID-19 disease. Ivermectin, a broad-spectrum anti-parasitic agent, has been shown to have anti-SARS-CoV-2 activity in Vero cells at a concentration of 5 {micro}M. These in vitro results triggered the investigation of ivermectin as a treatment option to alleviate COVID-19 disease. In April 2021, the World Health Organization stated, however, the following: "the current evidence on the use of ivermectin to treat COVID-19 patients is inconclusive". It is speculated that the in vivo concentration of ivermectin is too low to exert a strong antiviral effect. Here, we performed a head-to head comparison of the antiviral activity of ivermectin and a structurally related, but metabolically more stable, moxidectin in multiple in vitro models of SARS-CoV-2 infection, including physiologically relevant human respiratory epithelial cells. Both moxidectin and ivermectin exhibited antiviral activity in Vero E6 cells. Subsequent experiments revealed that the compounds predominantly act on a step after virus cell entry. Surprisingly, however, in human airway-derived cell models, moxidectin and ivermectin failed to inhibit SARS-CoV-2 infection, even at a concentration of 10 {micro}M. These disappointing results calls for a word of caution in the interpretation of anti-SARS-CoV-2 activity of drugs solely based on Vero cells. Altogether, these findings suggest that, even by using a high-dose regimen of ivermectin or switching to another drug in the same class are unlikely to be useful for treatment against SARS-CoV-2 in humans.

microbiology↗

bioRxiv2020

Resveratrol And Pterostilbene Potently Inhibit SARS-CoV-2 Infection In Vitro

The current COVID-19 pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has an enormous impact on human health and economy. In search for therapeutic options, researchers have proposed resveratrol, a food supplement with known antiviral, anti-inflammatory and anti-oxidant properties as an advantageous antiviral therapy for SARS-CoV-2 infection. Here, we provide evidence that both resveratrol and its metabolically more stable structural analog, pterostilbene, exhibit potent antiviral properties against SARS-CoV-2 in vitro. Resveratrol and pterostilbene showed antiviral activity in African green monkey kidney cells and in human primary bronchial epithelial cells cultured in an air-liquid interface system. Both compounds actively inhibit virus replication within infected cells as reduced virus progeny production was observed when the compound was added at post-inoculation conditions. Without replenishment of the compound, antiviral activity was observed up to roughly 5 rounds of replication, demonstrating the long-lasting effect of these compounds. Collectively, our data indicate that resveratrol and pterostilbene are promising antiviral compounds to treat SARS-CoV-2 infection. Because these results represent laboratory findings in cells, we advocate evaluation of these compounds in clinical trials before statements are made whether or not these drugs are advantageous for COVID-19 treatment.

microbiology↗

bioRxiv2020

Chikungunya virus requires an intact microtubule network for efficient viral genome delivery

Chikungunya virus (CHIKV) is a re-emerging mosquito-borne virus, which has rapidly spread around the globe thereby causing millions of infections. CHIKV is an enveloped virus belonging to the Togaviridae family and enters its host cell primarily via clathrin-mediated endocytosis. Upon internalization, the endocytic vesicle containing the virus particle moves through the cell and delivers the virus to early endosomes where membrane fusion is observed. Thereafter, the nucleocapsid dissociates and the viral RNA is translated into proteins. In this study, we examined the importance of the microtubule network during the early steps of infection and dissected the intracellular trafficking behavior of CHIKV particles during cell entry. We observed two distinct CHIKV intracellular trafficking patterns prior to membrane hemifusion. Whereas half of the CHIKV virions remained static during cell entry and fused in the cell periphery, the other half showed fast-directed microtubule-dependent movement prior to delivery to Rab5-positive early endosomes and predominantly fused in the perinuclear region of the cell. Disruption of the microtubule network reduced the number of infected cells. At these conditions, membrane hemifusion activity was not affected yet fusion was restricted to the cell periphery. Furthermore, follow-up experiments revealed that disruption of the microtubule network impairs the delivery of the viral genome to the cell cytosol. We therefore hypothesize that microtubules may direct the particle to a cellular location that is beneficial for establishing infection or aids in nucleocapsid uncoating. Author SummaryChikungunya virus (CHIKV) is an alphavirus that is transmitted to humans by infected mosquitoes. Disease symptoms can include fever, rash, myalgia, and long-lasting debilitating joint pains. Unfortunately, there is currently no licensed vaccine or antiviral treatment available to combat CHIKV. Understanding the virus:host interactions during the replication cycle of the virus is crucial for the development of effective antiviral therapies. In this study we elucidated the trafficking behavior of CHIKV particles early in infection. During cell entry, CHIKV virions require an intact microtubule network for efficient delivery of the viral genome into the host cell thereby increasing the chance to productively infect a cell.

microbiology↗

bioRxiv2020

Serotonergic drugs inhibit CHIKV infection at different stages of the cell entry pathway

Chikungunya virus (CHIKV) is an important re-emerging human pathogen transmitted by mosquitoes. The virus causes an acute febrile illness, chikungunya fever, which is characterized by headache, rash and debilitating (poly)arthralgia that can reside for months to years after infection. Currently, effective antiviral therapies and vaccines are lacking. Due to the high morbidity and economic burden in the countries affected by CHIKV, there is a strong need for new strategies to inhibit CHIKV replication. The serotonergic drug, 5-nonyloxytryptamine (5-NT), was previously identified as a potential host-directed inhibitor for CHIKV infection. In this study, we determined the mechanism of action by which the serotonin receptor agonist 5-NT controls CHIKV infection. Using time-of-addition and entry bypass assays we found that 5-NT predominantly inhibits CHIKV in the early phases of the replication cycle; at a step prior to RNA translation and genome replication. Intriguingly, however, no effect was seen during virus-cell binding, internalization, membrane fusion and gRNA release into the cell cytosol. Additionally, we show that the serotonin receptor antagonist MM also has antiviral properties towards CHIKV and specifically interferes with the cell entry process and/or membrane fusion. Taken together, pharmacological targeting of 5-HT receptors may represent a potent way to limit viral spread and disease severity. ImportanceThe rapid spread of mosquito-borne viral diseases in humans puts a huge economic burden on developing countries. For many of these infections, including Chikungunya virus (CHIKV), there are no specific treatment possibilities to alleviate disease symptoms. Understanding the virus:host interactions that are involved in the viral replication cycle is imperative for the rational design of therapeutic strategies. In this study, we discovered an antiviral compound and elucidated the mechanism of action and propose serotonergic drugs as potential host-directed antivirals for CHIKV.

microbiology↗

bioRxiv2020

Cooperative Chikungunya virus membrane fusion and its sub-stoichiometric inhibition by CHK-152 antibody

Chikungunya virus (CHIKV) presents a major burden on healthcare systems worldwide, but specific treatment remains unavailable. Attachment and fusion of CHIKV to the host cell membrane is mediated by the E1/E2 protein spikes. We used an in vitro single-particle fusion assay to study the effect of the potent, neutralizing antibody CHK-152 on CHIKV binding and fusion. We find that CHK-152 shields the virions, inhibiting interaction with the target membrane and inhibiting fusion. Analysis of the ratio of bound antibodies to epitopes implied that CHIKV fusion is a highly cooperative process. Further, dissociation of the antibody at lower pH results in a finely balanced kinetic competition between inhibition and fusion, suggesting a window of opportunity for the spike proteins to act and mediate fusion even in the presence of antibody.

microbiology↗