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Importance of charge interactions in Rift Valley fever virus attachment to host cells
Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi. (Magnus Evander)ORCID-id: 0000-0002-1269-4770
Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi, Virologi.
(Engelska)Manuskript (preprint) (Övrigt vetenskapligt)
Abstract [en]

The mosquito-borne Rift Valley fever virus (RVFV) cause disease in both humans and animals and can infect a large range of animals as well as humans. Many different cell types are infected both in vivo and in vitro. To enter a cell the virus needs to attach and enter, and this initial binding to the host cell surface could depend on both general mechanisms, and different specific receptors. Our aim was to characterize determinants for RVFV entry into its host cells.To examine RVFV attachment to host cells we based our experimental assay on RVF virus-like particles containing a reporter gene. The enveloped RVFV uses protruding glycoproteins (Gn and Gc) for attachment and entry and to investigate potential virus-cell surface interactions, the net surface charge of the glycoproteins was first calculated. The RVFV glycoprotein Gn had a predicted isoelectric point (pI) of 7.6 and a net positive charge of +6.9 at pH 7.0, suggesting a charge interaction between the Gn ectodomain and the negatively charged cell surface. RVFV Gc on the other hand, was highly negatively charged, -12.8 at neutral pH, most probably reflecting that Gc is not exposed until after receptor binding. To characterize the general conditions needed for RVFV attachment, cells or virus were treated with various compounds. Both sodium chloride and the negatively charged heparin inhibited RVF virus-like particle infection, strongly indicating that viral binding was charge-dependent. Treatment with sodium periodate pointed to a carbohydrate structure as a cellular interaction partner. Removal of sialic acid or heparan sulfate receptors on the cell surface by enzymatic treatment and blocking of the heparan sulfate receptor did not inhibit virus attachment.In conclusion, RVFV binding to host cells was charge dependent and the results point to a carbohydrate structure with negative charge as a potential attachment factor.

Nyckelord [en]
Rift Valley fever, receptor, entry, charge, carbohydrates
Nationell ämneskategori
Cell- och molekylärbiologi Mikrobiologi inom det medicinska området
Forskningsämne
molekylär medicin (medicinska vetenskaper); infektionssjukdomar
Identifikatorer
URN: urn:nbn:se:umu:diva-126473OAI: oai:DiVA.org:umu-126473DiVA, id: diva2:1033580
Tillgänglig från: 2016-10-07 Skapad: 2016-10-07 Senast uppdaterad: 2018-06-09
Ingår i avhandling
1. Rift Valley fever: consequences of virus-host interactions
Öppna denna publikation i ny flik eller fönster >>Rift Valley fever: consequences of virus-host interactions
2016 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

Rift Valley fever virus (RVFV) is a mosquito-borne virus which has the ability to infect a large variety of animals including humans in Africa and Arabian Peninsula. The abortion rate among these animals are close to 100%, and young animals develop severe disease which often are lethal.

In humans, Rift Valley fever (RVF) presents in most cases as a mild illness with influenza-like symptoms. However, in about 8% of the cases it progresses into a more severe disease with a high case fatality rate. Since there is such a high abortion rate among infected animals, a link between human miscarriage and RVFV has been suggested, but never proven.

We could in paper I for the first time show an association between acute RVFV infection and miscarriage in humans. We observed an increase in pregnant women arriving at the Port Sudan Hospital with fever of unknown origin, and several of the patients experienced miscarriage. When we analysed their blood samples for several viral diseases we found that many had an acute RVFV infection and of these, 54% experienced a miscarriage. The odds of having a miscarriage was 7 times higher for RVFV patients compared to the RVFV negative women of which only 12% miscarried. These results indicated that RVFV infection could be a contributing factor to miscarriage.

RVFV is an enveloped virus containing the viral glycoproteins n and c (Gn and Gc respectively), where Gn most likely is responsible for the initial cellular contact. The protein DC-SIGN on dendritic cells and the glycosaminoglycan heparan sulfate has been suggested as cellular receptors for RVFV, however other mechanisms are probably also involved in binding and entry. Charge is a driving force for molecular interaction and has been shown to be important for cellular attachment of several viruses, and in paper II we could show that when the charge around the cells was altered, the infection was affected. We also showed that Gn most likely has a positive charge at a physiological pH.

When we added negatively charged molecules to the viral particles before infection, we observed a decreased infection efficiency, which we also observed after removal of carbohydrate structures from the cell surface.

Our results suggested that the cellular interaction partner for initial attachment is a negatively charged carbohydrate. Further investigations into the mechanisms of RVFV cellular interactions has to be undertaken in order to understand, and ultimately prevent, infection and disease.

There is currently no vaccine approved for human use and no specific treatments for RVF, so there is a great need for developing safe effective drugs targeting this virus. We designed a whole-cell based high-throughput screen (HTS) assay which we used to screen libraries of small molecular compounds for anti-RVFV properties. After dose-response and toxicity analysis of the initial hits, we identified six safe and effective inhibitors of RVFV infection that with further testing could become drug candidates for treatment of RVF. This study demonstrated the application of HTS using a whole-cell virus replication reporter gene assay as an effective method to identify novel compounds with potential antiviral activity against RVFV.

Ort, förlag, år, upplaga, sidor
Umeå: Umeå universitet, 2016. s. 58
Serie
Umeå University medical dissertations, ISSN 0346-6612 ; 1843
Nyckelord
Rift Valley fever, Rift Valley fever virus, viral haemorrhagic fever, miscarriage, entry, charge, carbohydrates, high-throughput screening, antiviral, cell-based assay
Nationell ämneskategori
Infektionsmedicin Mikrobiologi inom det medicinska området
Forskningsämne
medicinsk virologi
Identifikatorer
urn:nbn:se:umu:diva-126602 (URN)978-91-7601-558-2 (ISBN)
Disputation
2016-11-04, Hörsal D, Unod T9, 9 trappor, Norrlands universitetssjukhus, Umeå, 09:00 (Engelska)
Opponent
Handledare
Tillgänglig från: 2016-10-14 Skapad: 2016-10-12 Senast uppdaterad: 2018-06-09Bibliografiskt granskad

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Baudin, MariaEvander, Magnus

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