Access through your institution Buy or subscribe Humankind faces regular outbreaks from emerging viruses, such as Ebola and the current SARS-CoV-2, or from viruses circulating at low levels

in the wild animal reservoir. Owing to the suddenness of virus expansion, the lack of knowledge of immune determinants and correlates of protection severely limit the ability to address the

outbreaks. Identified in 1976, Ebola virus is part of the _Filoviridae_ family and causes severe hemorrhagic fever with a mortality rate between 50% and 90%. From 2013 to 2020, more than

30,000 infections and 15,000 deaths have been reported worldwide. High antibody titers against Ebola virus GP are found in patients recovering from Ebola virus infection and correlate with

most of the T-cell responses mounted against Ebola virus were found to target NP.3 However, very few T-cell epitopes specific for both the Ebola GP and NP proteins have yet been identified

in humans.4,5 Only one study reported CD4 T-cell epitopes restricted to HLA-DR3.6 We therefore established a large-scale approach to identify CD4 T-cell epitopes using donors not exposed to

infection to anticipate the sudden rise of emerging viruses and applied this approach to the Ebola GP and NP proteins. Ten NP and eight GP peptides generated specific T-cell lines in at

least 50% of the tested donors. Interestingly, only a combination of four peptides (three from NP and one from GP) suffices to induce a T-cell response in all donors (Fig. S2) but

corresponds to only 15% of the total response. Alternatively, a combination of 18 peptides accounted for more than 50% of the total response (Fig. S2). We therefore extended the

characterization of the T cells raised against these peptides. Using HLA-specific antibodies, we found that most of the 18 T-cell epitopes are restricted to HLA-DR molecules, as predicted in

silico (Fig. 1c–e). Eight peptides were restricted at least partly to HLA-DP molecules, which shared HLA-DR common anchor residues, with the peptide NP80-99 being restricted to HLA-DP only.

We also assessed the T-cell cross-reactivity for the other Ebola strains (Fig. 1c–e). The initial peptide sequences were from the Ebola Zaire strain, as this strain is the first one

reported and is among the most virulent strains. Two other Ebola strains lethal to humans have been identified (Sudan and Bundibugyo), with the Reston strain being nonlethal in humans, and

there is only one nonfatal case reported (1994) with the Tai Forest strain (not considered here). The specificity of T-cell lines raised against Zaire epitopes was evaluated with

corresponding Sudan, Reston, and Bundibugyo peptides by IFN-γ ELISpot (Fig. 1c–e). Examples of partial and full cross-reactivity are shown in Fig. 1c and Fig. 1d, respectively. In addition

to three NP peptides, which were completely conserved across the Ebola strains, one NP peptide was completely conserved between the Sudan and Reston strains and presented 86%

cross-reactivity with Bundibugyo. Three NP peptides and two GP peptides exhibited a cross-reactivity greater than 66% to Sudan-derived peptides, which is the second most virulent Ebola

strain. Another peptide cross-reacted with the other strains, while 2 peptides cross-reacted with Bundibugyo only. Variants of three GP epitopes were not tested because they were too

divergent from their Zaire strain counterparts. We also investigated whether the identified T-cell epitopes were immunodominant and hence were naturally processed from the whole Ebola

proteins and presented to T cells. Peptide-specific T-cell lines were tested for their capacity to be activated by dendritic cells loaded with either the recombinant NP or GP Ebola protein

(Fig. 1c–e). Aside from those recognizing the peptide NP390-409, all the peptide-specific T-cell lines were stimulated by either the recombinant NP or GP protein. TCR V-Beta repertoire

analysis of 23 T-cell lines specific for 3 NP peptides (NP27-46, NP80-99 and NP147-166) and 8 T-cell lines specific for 3 GP peptides (GP31-50, GP60-79 and GP123-142) from 3 different donors

was conducted, showing that the T-cell response was shaped mainly by the individual repertoire of the donor (Table S2). This is a preview of subscription content, access via your

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* Log in * Learn about institutional subscriptions * Read our FAQs * Contact customer support REFERENCES * Wong, G., Kobinger, G. P. & Qiu, X. Characterization of host immune responses

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_Hum. Vaccines Immunother._ 13, 2824–2836 (2017). Article  Google Scholar  * Kwok, W. W. et al. Frequency of epitope-specific naive CD4+ T cells correlates with immunodominance in the human

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diversity and response magnitude. _Immunity_ 27, 203–213 (2007). Article  CAS  Google Scholar  Download references ACKNOWLEDGEMENTS The research leading to these results was supported by the

Innovative Medicines Initiative Joint Undertaking PEVIA project under grant agreement #116088, the resources of which comprise financial contributions from the European Union. The authors

thank Tiphanie Pruvost and Evelyne Correia from SIMoS for helpful discussions and technical advice. AUTHOR INFORMATION AUTHORS AND AFFILIATIONS * Département Médicament et Technologie pour

la Santé, SIMoS, Université Paris-Saclay, CEA-Saclay, 91191, Gif surYvette, France Yann Gallais, Raphaël Sierocki, Gautier Lhomme, Coline Sivelle & Bernard Maillère * Excellgene,

Monthey, Switzerland Divor Kiseljak & Florian Wurm * Wiratech Europe, Genopole, 91000, Evry, France Sami Djoulah * Vaxeal holding SA, Vevey, Switzerland Ahmed Bouzidi & Jérôme

Kerzerho Authors * Yann Gallais View author publications You can also search for this author inPubMed Google Scholar * Raphaël Sierocki View author publications You can also search for this

author inPubMed Google Scholar * Gautier Lhomme View author publications You can also search for this author inPubMed Google Scholar * Coline Sivelle View author publications You can also

search for this author inPubMed Google Scholar * Divor Kiseljak View author publications You can also search for this author inPubMed Google Scholar * Florian Wurm View author publications

You can also search for this author inPubMed Google Scholar * Sami Djoulah View author publications You can also search for this author inPubMed Google Scholar * Ahmed Bouzidi View author

publications You can also search for this author inPubMed Google Scholar * Jérôme Kerzerho View author publications You can also search for this author inPubMed Google Scholar * Bernard

Maillère View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS Y.G., B.M., A.B, J.K., S.D., and F.W. designed the experiments; Y.G., R.S., G.L.,

C.S, and D.K. performed the experiments; Y.G., G.L., and B.M. analyzed the data; and Y.G. and B.M. wrote the paper. CORRESPONDING AUTHOR Correspondence to Bernard Maillère. ETHICS

DECLARATIONS COMPETING INTERESTS Y.G. and B.M. are inventors of a pending patent. SUPPLEMENTARY INFORMATION SUPPLEMENTAL MATERIAL RIGHTS AND PERMISSIONS Reprints and permissions ABOUT THIS

ARTICLE CITE THIS ARTICLE Gallais, Y., Sierocki, R., Lhomme, G. _et al._ Large-scale mapping of the Ebola NP and GP proteins reveals multiple immunoprevalent and conserved CD4 T-cell

epitopes. _Cell Mol Immunol_ 18, 1323–1325 (2021). https://doi.org/10.1038/s41423-020-0455-2 Download citation * Received: 14 April 2020 * Accepted: 18 April 2020 * Published: 12 May 2020 *

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