Usama Ashraf

All Publications

• Durable protection against the SARS-CoV-2 Omicron variant is induced by an adjuvanted subunit vaccine. Science translational medicine Arunachalam, P. S., Feng, Y., Ashraf, U., Hu, M., Walls, A. C., Edara, V. V., Zarnitsyna, V. I., Aye, P. P., Golden, N., Miranda, M. C., Green, K. W., Threeton, B. M., Maness, N. J., Beddingfield, B. J., Bohm, R. P., Scheuermann, S. E., Goff, K., Dufour, J., Russell-Lodrigue, K., Kepl, E., Fiala, B., Wrenn, S., Ravichandran, R., Ellis, D., Carter, L., Rogers, K., Shirreff, L. M., Ferrell, D. E., Deb Adhikary, N. R., Fontenot, J., Hammond, H. L., Frieman, M., Grifoni, A., Sette, A., O'Hagan, D. T., Van Der Most, R., Rappuoli, R., Villinger, F., Kleanthous, H., Rappaport, J., Suthar, M. S., Veesler, D., Wang, T. T., King, N. P., Pulendran, B. 2022; 14 (658): eabq4130

  Abstract

  Despite the remarkable efficacy of COVID-19 vaccines, waning immunity and the emergence of SARS-CoV-2 variants such as Omicron represents a global health challenge. Here, we present data from a study in nonhuman primates demonstrating durable protection against the Omicron BA.1 variant induced by a subunit SARS-CoV-2 vaccine comprising the receptor binding domain of the ancestral strain (RBD-Wu) on the I53-50 nanoparticle adjuvanted with AS03, which was recently authorized for use in individuals 18 years or older. Vaccination induced neutralizing antibody (nAb) titers that were maintained at high concentrations for at least 1 year after two doses, with a pseudovirus nAb geometric mean titer (GMT) of 1978 and a live virus nAb GMT of 1331 against the ancestral strain but not against the Omicron BA.1 variant. However, a booster dose at 6 to 12 months with RBD-Wu or RBD-beta (RBD from the Beta variant) displayed on I53-50 elicited high neutralizing titers against the ancestral and Omicron variants. In addition, we observed persistent neutralization titers against a panel of sarbecoviruses, including SARS-CoV. Furthermore, there were substantial and persistent memory T and B cell responses reactive to Beta and Omicron variants. Vaccination resulted in protection against Omicron infection in the lung and suppression of viral burden in the nares at 6 weeks after the final booster immunization. Even at 6 months after vaccination, we observed protection in the lung and rapid control of virus in the nares. These results highlight the durable and cross-protective immunity elicited by the AS03-adjuvanted RBD-I53-50 nanoparticle vaccine.

  View details for DOI 10.1126/scitranslmed.abq4130

  View details for PubMedID 35976993

• Zika virus causes placental pyroptosis and associated adverse fetal outcomes by activating GSDME ELIFE Zhao, Z., Li, Q., Ashraf, U., Yang, M., Zhu, W., Gu, J., Chen, Z., Gu, C., Si, Y., Cao, S., Ye, J., Carette, J. E. 2022; 11

  Abstract

  Zika virus (ZIKV) can be transmitted from mother to fetus during pregnancy, causing adverse fetal outcomes. Several studies have indicated that ZIKV can damage the fetal brain directly; however, whether the ZIKV-induced maternal placental injury contributes to adverse fetal outcomes is sparsely defined. Here, we demonstrated that ZIKV causes the pyroptosis of placental cells by activating the executor gasdermin E (GSDME) in vitro and in vivo. Mechanistically, TNF-α release is induced upon the recognition of viral genomic RNA by RIG-I, followed by activation of caspase-8 and caspase-3 to ultimately escalate the GSDME cleavage. Further analyses revealed that the ablation of GSDME or treatment with TNF-α receptor antagonist in ZIKV-infected pregnant mice attenuates placental pyroptosis, which consequently confers protection against adverse fetal outcomes. In conclusion, our study unveils a novel mechanism of ZIKV-induced adverse fetal outcomes via causing placental cell pyroptosis, which provides new clues for developing therapies for ZIKV-associated diseases.

  View details for DOI 10.7554/eLife.73792

  View details for Web of Science ID 000841489100001

  View details for PubMedID 35972780

  View details for PubMedCentralID PMC9381041

• Early non-neutralizing, afucosylated antibody responses are associated with COVID-19 severity. Science translational medicine Chakraborty, S., Gonzalez, J. C., Sievers, B. L., Mallajosyula, V., Chakraborty, S., Dubey, M., Ashraf, U., Cheng, B. Y., Kathale, N., Tran, K. Q., Scallan, C., Sinnott, A., Cassidy, A., Chen, S. T., Gelbart, T., Gao, F., Golan, Y., Ji, X., Kim-Schulze, S., Prahl, M., Gaw, S. L., Gnjatic, S., Marron, T. U., Merad, M., Arunachalam, P. S., Boyd, S. D., Davis, M. M., Holubar, M., Khosla, C., Maecker, H. T., Maldonado, Y., Mellins, E. D., Nadeau, K. C., Pulendran, B., Singh, U., Subramanian, A., Utz, P. J., Sherwood, R., Zhang, S., Jagannathan, P., Tan, G. S., Wang, T. T. 1800: eabm7853

  Abstract

  A damaging inflammatory response is implicated in the pathogenesis of severe coronavirus disease 2019 (COVID-19), but mechanisms contributing to this response are unclear. In two prospective cohorts, early non-neutralizing, afucosylated IgG antibodies specific to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were associated with progression from mild to more severe COVID-19. In contrast to the antibody structures that were associated with disease progression, antibodies that were elicited by mRNA SARS-CoV-2 vaccines were instead highly fucosylated and enriched in sialylation, both modifications that reduce the inflammatory potential of IgG. To study the biology afucosylated IgG immune complexes, we developed an in vivo model that revealed that human IgG-Fc gamma receptor (FcgammaR) interactions could regulate inflammation in the lung. Afucosylated IgG immune complexes isolated from COVID-19 patients induced inflammatory cytokine production and robust infiltration of the lung by immune cells. By contrast, vaccine-elicited IgG did not promote an inflammatory lung response. Together, these results show that IgG-FcgammaR interactions are able to regulate inflammation in the lung and may define distinct lung activities associated with the IgG that are associated with severe COVID-19 and protection against infection with SARS-CoV-2.

  View details for DOI 10.1126/scitranslmed.abm7853

  View details for PubMedID 35040666

• Pathogenicity and virulence of Japanese encephalitis virus: Neuroinflammation and neuronal cell damage. Virulence Ashraf, U., Ding, Z., Deng, S., Ye, J., Cao, S., Chen, Z. 2021; 12 (1): 968-980

  Abstract

  Thousands of human deaths occur annually due to Japanese encephalitis (JE), caused by Japanese encephalitis virus. During the virus infection of the central nervous system, reactive gliosis, uncontrolled inflammatory response, and neuronal cell death are considered as the characteristic features of JE. To date, no specific treatment has been approved to overcome JE, indicating a need for the development of novel therapies. In this article, we focused on basic biological mechanisms in glial (microglia and astrocytes) and neuronal cells that contribute to the onset of neuroinflammation and neuronal cell damage during Japanese encephalitis virus infection. We also provided comprehensive knowledge about anti-JE therapies tested in clinical or pre-clinical settings, and discussed recent therapeutic strategies that could be employed for JE treatment. The improved understanding of JE pathogenesis might lay a foundation for the development of novel therapies to halt JE.Abbreviations AKT: a serine/threonine-specific protein kinase; AP1: activator protein 1; ASC: apoptosis-associated speck-like protein containing a CARD; ASK1: apoptosis signal-regulated kinase 1; ATF3/4/6: activating transcription factor 3/4/6; ATG5/7: autophagy-related 5/7; BBB: blood-brain barrier; Bcl-3/6: B-cell lymphoma 3/6 protein; CCL: C-C motif chemokine ligand; CCR2: C-C motif chemokine receptor 2; CHOP: C/EBP homologous protein; circRNA: circular RNA; CNS: central nervous system; CXCL: C-X-C motif chemokine ligand; dsRNA: double-stranded RNA; EDEM1: endoplasmic reticulum degradation enhancer mannosidase alpha-like 1; eIF2-ɑ: eukaryotic initiation factor 2 alpha; ER: endoplasmic reticulum; ERK: extracellular signal-regulated kinase; GRP78: 78-kDa glucose-regulated protein; ICAM: intercellular adhesion molecule; IFN: interferon; IL: interleukin; iNOS: inducible nitric oxide synthase; IRAK1/2: interleukin-1 receptor-associated kinase 1/2; IRE-1: inositol-requiring enzyme 1; IRF: interferon regulatory factor; ISG15: interferon-stimulated gene 15; JE: Japanese encephalitis; JEV: Japanese encephalitis virus; JNK: c-Jun N-terminal kinase; LAMP2: lysosome-associated membrane protein type 2; LC3-I/II: microtubule-associated protein 1 light chain 3-I/II; lncRNA: long non-coding RNA; MAPK: mitogen-activated protein kinase; miR/miRNA: microRNA; MK2: mitogen-activated protein kinase-activated protein kinase 2; MKK4: mitogen-activated protein kinase kinase 4; MLKL: mixed-linage kinase domain-like protein; MMP: matrix metalloproteinase; MyD88: myeloid differentiation factor 88; Nedd4: neural precursor cell-expressed developmentally downregulated 4; NF-κB: nuclear factor kappa B; NKRF: nuclear factor kappa B repressing factor; NLRP3: NLR family pyrin domain containing 3; NMDAR: N-methyl-D-aspartate receptor; NO: nitric oxide; NS2B/3/4: JEV non-structural protein 2B/3/4; P: phosphorylation. p38: mitogen-activated protein kinase p38; PKA: protein kinase A; PAK4: p21-activated kinase 4; PDFGR: platelet-derived growth factor receptor; PERK: protein kinase R-like endoplasmic reticulum kinase; PI3K: phosphoinositide 3-kinase; PTEN: phosphatase and tensin homolog; Rab7: Ras-related GTPase 7; Raf: proto-oncogene tyrosine-protein kinase Raf; Ras: a GTPase; RIDD: regulated IRE-1-dependent decay; RIG-I: retinoic acid-inducible gene I; RIPK1/3: receptor-interacting protein kinase 1/3; RNF11/125: RING finger protein 11/125; ROS: reactive oxygen species; SHIP1: SH2-containing inositol 5' phosphatase 1; SOCS5: suppressor of cytokine signaling 5; Src: proto-oncogene tyrosine-protein kinase Src; ssRNA = single-stranded RNA; STAT: signal transducer and activator of transcription; TLR: toll-like receptor; TNFAIP3: tumor necrosis factor alpha-induced protein 3; TNFAR: tumor necrosis factor alpha receptor; TNF-α: tumor necrosis factor-alpha; TRAF6: tumor necrosis factor receptor-associated factor 6; TRIF: TIR-domain-containing adapter-inducing interferon-β; TRIM25: tripartite motif-containing 25; VCAM: vascular cell adhesion molecule; ZO-1: zonula occludens-1.

  View details for DOI 10.1080/21505594.2021.1899674

  View details for PubMedID 33724154

  View details for PubMedCentralID PMC7971234

• Influenza virus infection induces widespread alterations of host cell splicing. NAR genomics and bioinformatics Ashraf, U., Benoit-Pilven, C., Navratil, V., Ligneau, C., Fournier, G., Munier, S., Sismeiro, O., Coppée, J. Y., Lacroix, V., Naffakh, N. 2020; 2 (4): lqaa095

  Abstract

  Influenza A viruses (IAVs) use diverse mechanisms to interfere with cellular gene expression. Although many RNA-seq studies have documented IAV-induced changes in host mRNA abundance, few were designed to allow an accurate quantification of changes in host mRNA splicing. Here, we show that IAV infection of human lung cells induces widespread alterations of cellular splicing, with an overall increase in exon inclusion and decrease in intron retention. Over half of the mRNAs that show differential splicing undergo no significant changes in abundance or in their 3' end termination site, suggesting that IAVs can specifically manipulate cellular splicing. Among a randomly selected subset of 21 IAV-sensitive alternative splicing events, most are specific to IAV infection as they are not observed upon infection with VSV, induction of interferon expression or induction of an osmotic stress. Finally, the analysis of splicing changes in RED-depleted cells reveals a limited but significant overlap with the splicing changes in IAV-infected cells. This observation suggests that hijacking of RED by IAVs to promote splicing of the abundant viral NS1 mRNAs could partially divert RED from its target mRNAs. All our RNA-seq datasets and analyses are made accessible for browsing through a user-friendly Shiny interface (http://virhostnet.prabi.fr:3838/shinyapps/flu-splicing or https://github.com/cbenoitp/flu-splicing).

  View details for DOI 10.1093/nargab/lqaa095

  View details for PubMedID 33575639

  View details for PubMedCentralID PMC7680258

• Destabilization of the human RED-SMU1 splicing complex as a basis for host-directed antiinfluenza strategy. Proceedings of the National Academy of Sciences of the United States of America Ashraf, U., Tengo, L., Le Corre, L., Fournier, G., Busca, P., McCarthy, A. A., Rameix-Welti, M. A., Gravier-Pelletier, C., Ruigrok, R. W., Jacob, Y., Vidalain, P. O., Pietrancosta, N., Crépin, T., Naffakh, N. 2019; 116 (22): 10968-10977

  Abstract

  New therapeutic strategies targeting influenza are actively sought due to limitations in current drugs available. Host-directed therapy is an emerging concept to target host functions involved in pathogen life cycles and/or pathogenesis, rather than pathogen components themselves. From this perspective, we focused on an essential host partner of influenza viruses, the RED-SMU1 splicing complex. Here, we identified two synthetic molecules targeting an α-helix/groove interface essential for RED-SMU1 complex assembly. We solved the structure of the SMU1 N-terminal domain in complex with RED or bound to one of the molecules identified to disrupt this complex. We show that these compounds inhibiting RED-SMU1 interaction also decrease endogenous RED-SMU1 levels and inhibit viral mRNA splicing and viral multiplication, while preserving cell viability. Overall, our data demonstrate the potential of RED-SMU1 destabilizing molecules as an antiviral therapy that could be active against a wide range of influenza viruses and be less prone to drug resistance.

  View details for DOI 10.1073/pnas.1901214116

  View details for PubMedID 31076555

  View details for PubMedCentralID PMC6561211

• Advances in Analyzing Virus-Induced Alterations of Host Cell Splicing. Trends in microbiology Ashraf, U., Benoit-Pilven, C., Lacroix, V., Navratil, V., Naffakh, N. 2019; 27 (3): 268-281

  Abstract

  Alteration of host cell splicing is a common feature of many viral infections which is underappreciated because of the complexity and technical difficulty of studying alternative splicing (AS) regulation. Recent advances in RNA sequencing technologies revealed that up to several hundreds of host genes can show altered mRNA splicing upon viral infection. The observed changes in AS events can be either a direct consequence of viral manipulation of the host splicing machinery or result indirectly from the virus-induced innate immune response or cellular damage. Analysis at a higher resolution with single-cell RNAseq, and at a higher scale with the integration of multiple omics data sets in a systems biology perspective, will be needed to further comprehend this complex facet of virus-host interactions.

  View details for DOI 10.1016/j.tim.2018.11.004

  View details for PubMedID 30577974

• MicroRNA-22 negatively regulates poly(I:C)-triggered type I interferon and inflammatory cytokine production via targeting mitochondrial antiviral signaling protein (MAVS). Oncotarget Wan, S., Ashraf, U., Ye, J., Duan, X., Zohaib, A., Wang, W., Chen, Z., Zhu, B., Li, Y., Chen, H., Cao, S. 2016; 7 (47): 76667-76683

  Abstract

  MicroRNAs (miRNAs) are small non-coding RNAs that play important roles in regulating the host immune response. Here we found that miR-22 is induced in glial cells upon stimulation with poly(I:C). Overexpression of miR-22 in the cultured cells resulted in decreased activity of interferon regulatory factor-3 and nuclear factor-kappa B, which in turn led to reduced expression of interferon-β and inflammatory cytokines, including tumor necrosis factor-α, interleukin-1β, interleukin-6, and chemokine (C-C motif) ligand 5, upon stimulation with poly(I:C), whereas knockdown of miR-22 had the opposite effect. We used a combination of bioinformatics and experimental techniques to demonstrate that mitochondrial antiviral signaling protein (MAVS), which positively regulates type I interferon production, is a novel target of miR-22. Overexpression of miR-22 decreased the activity of a luciferase reporter containing the MAVS 3'-untranslated region and led to decreased MAVS mRNA and protein levels. In contrast, ectopic expression of miR-22 inhibitor led to elevated MAVS expression. Collectively, our results demonstrate that miR-22 negatively regulates poly(I:C)-induced production of type I interferon and inflammatory cytokines via targeting MAVS.

  View details for DOI 10.18632/oncotarget.12395

  View details for PubMedID 27705941

  View details for PubMedCentralID PMC5363539

• MicroRNA-19b-3p Modulates Japanese Encephalitis Virus-Mediated Inflammation via Targeting RNF11. Journal of virology Ashraf, U., Zhu, B., Ye, J., Wan, S., Nie, Y., Chen, Z., Cui, M., Wang, C., Duan, X., Zhang, H., Chen, H., Cao, S. 2016; 90 (9): 4780-4795

  Abstract

  Japanese encephalitis virus (JEV) can invade the central nervous system and consequently induce neuroinflammation, which is characterized by profound neuronal cell damage accompanied by astrogliosis and microgliosis. Albeit microRNAs (miRNAs) have emerged as major regulatory noncoding RNAs with profound effects on inflammatory response, it is unknown how astrocytic miRNAs regulate JEV-induced inflammation. Here, we found the involvement of miR-19b-3p in regulating the JEV-induced inflammatory responsein vitroandin vivo The data demonstrated that miR-19b-3p is upregulated in cultured cells and mouse brain tissues during JEV infection. Overexpression of miR-19b-3p led to increased production of inflammatory cytokines, including tumor necrosis factor alpha, interleukin-6, interleukin-1β, and chemokine (C-C motif) ligand 5, after JEV infection, whereas knockdown of miR-19b-3p had completely opposite effects. Mechanistically, miR-19b-3p modulated the JEV-induced inflammatory response via targeting ring finger protein 11, a negative regulator of nuclear factor kappa B signaling. We also found that inhibition of ring finger protein 11 by miR-19b-3p resulted in accumulation of nuclear factor kappa B in the nucleus, which in turn led to higher production of inflammatory cytokines.In vivosilencing of miR-19b-3p by a specific antagomir reinvigorates the expression level of RNF11, which in turn reduces the production of inflammatory cytokines, abrogates gliosis and neuronal cell death, and eventually improves the survival rate in the mouse model. Collectively, our results demonstrate that miR-19b-3p positively regulates the JEV-induced inflammatory response. Thus, miR-19b-3p targeting may constitute a thought-provoking approach to rein in JEV-induced inflammation.Japanese encephalitis virus (JEV) is one of the major causes of acute encephalitis in humans worldwide. The pathological features of JEV-induced encephalitis are inflammatory reactions and neurological diseases resulting from glia activation. MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression posttranscriptionally. Accumulating data indicate that miRNAs regulate a variety of cellular processes, including the host inflammatory response under pathological conditions. Recently, a few studies demonstrated the role of miRNAs in a JEV-induced inflammatory response in microglia; however, their role in an astrocyte-derived inflammatory response is largely unknown. The present study reveals that miR-19b-3p targets ring finger protein 11 in glia and promotes inflammatory cytokine production by enhancing nuclear factor kappa B activity in these cells. Moreover, administration of an miR-19b-3p-specific antagomir in JEV-infected mice reduces neuroinflammation and lethality. These findings suggest a new insight into the molecular mechanism of the JEV-induced inflammatory response and provide a possible therapeutic entry point for treating viral encephalitis.

  View details for DOI 10.1128/JVI.02586-15

  View details for PubMedID 26937036

  View details for PubMedCentralID PMC4836334

• Spring viraemia of carp virus: recent advances. The Journal of general virology Ashraf, U., Lu, Y., Lin, L., Yuan, J., Wang, M., Liu, X. 2016; 97 (5): 1037-1051

  Abstract

  Spring viraemia of carp is an environmentally and economically important disease affecting cyprinids, primarily common carp (Cyprinus carpio). The causative agent of this disease is Spring viraemia of carp virus (SVCV) - a member of the genus Vesiculovirus of the family Rhabdoviridae. The disease is presently endemic in Europe, America and several Asian countries, where it causes significant morbidity and mortality in affected fish. SVCV infection is generally associated with exophthalmia; abdominal distension; petechial haemorrhage of the skin, gills, eyes and internal organs; degeneration of the gill lamellae; a swollen and coarse-textured spleen; hepatic necrosis; enteritis; and pericarditis. The SVCV genome is composed of linear, negative-sense, ssRNA containing five genes in the order 3'-N-P-M-G-L-5', encoding a nucleoprotein, phosphoprotein, matrix protein, glycoprotein and RNA-dependent RNA polymerase, respectively. Fully sequenced SVCV strains exhibit distinct amino acid substitutions at unique positions, which may contribute to as-yet unknown strain-specific characteristics. To advance the study of SVCV and the control of spring viraemia of carp disease in the future, this review summarizes our current understanding of SVCV in terms of its genomic characteristics, genetic diversity and pathogenesis, and provides insights into antiviral immunity against SVCV, diagnosis of SVCV and vaccination strategies to combat SVCV.

  View details for DOI 10.1099/jgv.0.000436

  View details for PubMedID 26905065

• Usutu virus: an emerging flavivirus in Europe. Viruses Ashraf, U., Ye, J., Ruan, X., Wan, S., Zhu, B., Cao, S. 2015; 7 (1): 219-38

  Abstract

  Usutu virus (USUV) is an African mosquito-borne flavivirus belonging to the Japanese encephalitis virus serocomplex. USUV is closely related to Murray Valley encephalitis virus, Japanese encephalitis virus, and West Nile virus. USUV was discovered in South Africa in 1959. In Europe, the first true demonstration of circulation of USUV was reported in Austria in 2001 with a significant die-off of Eurasian blackbirds. In the subsequent years, USUV expanded to neighboring countries, including Italy, Germany, Spain, Hungary, Switzerland, Poland, England, Czech Republic, Greece, and Belgium, where it caused unusual mortality in birds. In 2009, the first two human cases of USUV infection in Europe have been reported in Italy, causing meningoencephalitis in immunocompromised patients. This review describes USUV in terms of its life cycle, USUV surveillance from Africa to Europe, human cases, its cellular tropism and pathogenesis, its genetic relationship with other flaviviruses, genetic diversity among USUV strains, its diagnosis, and a discussion of the potential future threat to Asian countries.

  View details for DOI 10.3390/v7010219

  View details for PubMedID 25606971

  View details for PubMedCentralID PMC4306835