Antigenic Characterization of the HCMV gH/gL/gO and Pentamer Cell Entry Complexes Reveals Binding Sites for Potently Neutralizing Human Antibodies

Ciferri, Claudio; Chandramouli, Sumana; Leitner, Alexander; Donnarumma, Danilo; Cianfrocco, Michael A.; Gerrein, Rachel; Friedrich, Kristian; Aggarwal, Yukti; Palladino, Giuseppe; Aebersold, Ruedi; Norais, Nathalie; Settembre, Ethan C.; Carfı́, Andrea

doi:10.1371/journal.ppat.1005230

Cited by 71 publications

(70 citation statements)

References 47 publications

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“…This analysis revealed that the gH/UL116 complex has a boot-shaped structure similar to that of gH/gL. 3G16 and MSL-109 Fabs bind to the C-terminal domain and to the "heel" of the gH molecule, respectively, as previously described for gH/gL (54,72). Furthermore, the EM analysis and binding data (unpublished results) suggest that UL116 occupies a position similar to that of gL at the N-terminal end of gH (Fig.…”

Section: Resultssupporting

confidence: 69%

“…We previously used single-particle negative-stain EM to characterize gH/gL, gH/gL/gO, and Pentamer complexes bound to neutralizing antibodies (54,72). Our data revealed that HCMV gH/gL has a boot-shaped structure similar to that of HSV-2 gH/gL (73) and that gO and the UL128/UL130/UL131A subunits bind to the N-terminal end of gH/gL in gH/gL/gO and Pentamer, respectively.…”

Section: Resultsmentioning

confidence: 82%

“…Here, we extended the EM analysis to the purified gH/ UL116 complex bound to either MSL-109 or 3G16 antibody (41,72). The trimeric gH/UL116/3G16 and gH/UL116/MSL-109 complexes were analyzed by negative-stain EM and single-particle analysis.…”

Section: Resultsmentioning

confidence: 99%

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The Human Cytomegalovirus UL116 Gene Encodes an Envelope Glycoprotein Forming a Complex with gH Independently from gL

Caló

Cortese

Ciferri³

et al. 2016

J Virol

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Human cytomegalovirus (HCMV) is a major cause of morbidity and mortality in transplant patients and is the leading viral cause of birth defects after congenital infection. HCMV infection relies on the recognition of cell-specific receptors by one of the viral envelope glycoprotein complexes. Either the gH/gL/gO or the gH/gL/UL128/UL130/UL131A (Pentamer) complex has been found to fulfill this role, accounting for HCMV entry into almost all cell types. We have studied the UL116 gene product, a putative open reading frame identified by in silico analysis and predicted to code for a secreted protein. Virus infection experiments in mammalian cells demonstrated that UL116 is expressed late in the HCMV replication cycle and is a heavily glycosylated protein that first localizes to the cellular site of virus assembly and then inserts into the virion envelope. Transient-transfection studies revealed that UL116 is efficiently transported to the plasma membrane when coexpressed with gH and that gL competes with UL116 for gH binding. Further evidence for gH/UL116 complex formation was obtained by coimmunoprecipitation experiments on both transfected and infected cells and biochemical characterization of the purified complex. In summary, our results show that the product of the UL116 gene is an HCMV envelope glycoprotein that forms a novel gH-based complex alternative to gH/gL. Remarkably, the gH/UL116 complex is the first herpesvirus gH-based gL-less complex. IMPORTANCE HCMV infection can cause severe disease in immunocompromised adults and infants infected in utero.The dissection of the HCMV entry machinery is important to understand the mechanism of viral infection and to identify new vaccine antigens. The gH/gL/gO and gH/gL/UL128/UL130/UL131 (Pentamer) complexes play a key role in HCMV cell entry and tropism. Both complexes are formed by an invariant gH/gL scaffold on which the other subunits assemble. Here, we show that the UL116 gene product is expressed in infected cells and forms a heterodimer with gH. The gH/UL116 complex is carried on the infectious virions, although in smaller amounts than gH/gL complexes. No gH/UL116/gL ternary complex formed in transfected cells, suggesting that the gH/UL116 complex is independent from gL. This new gH-based gL-free complex represents a potential target for a protective HCMV vaccine and opens new perspectives on the comprehension of the HCMV cell entry mechanism and tropism. H uman cytomegalovirus (HCMV) is a ubiquitous betaherpesvirus infecting 40 to 60% of the human population (1).HCMV infection usually is mild or asymptomatic in immunocompetent individuals, while it can cause severe disease in immunocompromised adults. Infants infected in utero can suffer from disseminated HCMV disease and impaired neurological development (2). Transplant patients do not adequately control HCMV infection or reactivation, often resulting in graft rejection and death (3). Preventing primary HCMV infection could significantly decrease the frequency of organ rejection in seronegative recip...

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Section: Resultssupporting

confidence: 69%

Section: Resultsmentioning

confidence: 82%

See 1 more Smart Citation

The Human Cytomegalovirus UL116 Gene Encodes an Envelope Glycoprotein Forming a Complex with gH Independently from gL

Caló

Cortese

Ciferri³

et al. 2016

J Virol

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“…Possible explanations include a major role played by memory B cells of donor and recipient origins [122]. The gH/gL-pentamer complex represents the platform for prophylactic Pentamer-based vaccines of absolute interest [123,124].…”

Section: Vaccinesmentioning

confidence: 99%

Treatment of CMV infection after allogeneic hematopoietic stem cell transplantation

Madon

Giaccone

Festuccia

et al. 2016

Expert Review of Hematology

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“…Previous studies demonstrated that the PC comprises predominantly quaternary conformational epitopes that are formed by two or more subunits and in only rare cases epitopes that are constituted by single subunits of the PC (28,29). In addition, Ciferri et al identified amino acid residues that are either in close proximity to or part of conformational neutralizing epitopes within the PC (42). However, amino acid residues that constitute the minimal target sequence of a potent PC-specific NAb have not been described.…”

Section: Discussionmentioning

confidence: 99%

Identification of a Continuous Neutralizing Epitope within UL128 of Human Cytomegalovirus

Chiuppesi

Kaltcheva

Zhao

et al. 2017

J Virol

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As human cytomegalovirus (HCMV) is the most common infectious cause of fetal anomalies during pregnancy, development of a vaccine that prevents HCMV infection is considered a global health priority. Although HCMV immune correlates of protection are only poorly defined, neutralizing antibodies (NAb) targeting the envelope pentamer complex (PC) composed of the subunits gH, gL, UL128, UL130, and UL131A are thought to contribute to the prevention of HCMV infection. Here, we describe a continuous target sequence within UL128 that is recognized by a previously isolated potent PC-specific NAb termed 13B5. By using peptide-based scanning procedures, we identified a 13-amino-acid-long target sequence at the UL128 C terminus that binds the 13B5 antibody with an affinity similar to that of the purified PC. In addition, the 13B5 binding site is universally conserved in HCMV, contains a previously described UL128/gL interaction site, and interferes with the 13B5 neutralizing function, indicating that the 13B5 epitope sequence is located within the PC at a site of critical importance for HCMV neutralization. Vaccination of mice with peptides containing the 13B5 target sequence resulted in the robust stimulation of binding antibodies and, in a subset of immunized animals, in the induction of detectable NAb, supporting that the identified 13B5 target sequence constitutes a PC-specific neutralizing epitope. These findings provide evidence for the discovery of a continuous neutralizing epitope within the UL128 subunit of the PC that could be an important target of humoral immune responses that are involved in protection against congenital HCMV infection.IMPORTANCE Neutralizing antibodies (NAb) targeting the human cytomegalovirus (HCMV) envelope pentamer complex (PC) are thought to be important for preventing HCMV transmission from the mother to the fetus, thereby mitigating severe developmental disabilities in newborns. However, the epitope sequences within the PC that are recognized by these potentially protective antibody responses are only poorly defined. Here, we provide evidence for the existence of a highly conserved, continuous, PC-specific epitope sequence that appears to be located within the PC at a subunit interaction site of critical importance for HCMV neutralization. These discoveries provide insights into a continuous PC-specific neutralizing epitope, which could be an important target for a vaccine formulation to interfere with congenital HCMV infection.

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Antigenic Characterization of the HCMV gH/gL/gO and Pentamer Cell Entry Complexes Reveals Binding Sites for Potently Neutralizing Human Antibodies

Cited by 71 publications

References 47 publications

The Human Cytomegalovirus UL116 Gene Encodes an Envelope Glycoprotein Forming a Complex with gH Independently from gL

The Human Cytomegalovirus UL116 Gene Encodes an Envelope Glycoprotein Forming a Complex with gH Independently from gL

Treatment of CMV infection after allogeneic hematopoietic stem cell transplantation

Identification of a Continuous Neutralizing Epitope within UL128 of Human Cytomegalovirus

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