Integrin α<sub>v</sub>β<sub>3</sub> mediates rotavirus cell entry

C Guerrero

E Méndez

S Zárate

P Isa

S López

C Arias

Departamento de Genética y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos 62250, Mexico

^{Permanent address: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional de Colombia, Bogotá, Colombia.}

^{To whom reprint requests should be addressed at: Instituto de Biotecnología/UNAM, A.P. 510–3, Colonia Miraval, Cuernavaca, Morelos 62250, Mexico. E-mail:}xm.manu.tbi@saira.

Edited by Wolfgang K. Joklik, Duke University Medical Center, Durham, NC, and approved October 4, 2000

Received 2000 Jun 28

Abstract

Rotavirus strains differ in their need for sialic acid (SA) for initial binding to the cell surface; however, the existence of a postattachment cell receptor, common to most, if not all, rotavirus strains, has been proposed. In the present study, antibodies to the α_v and β₃ integrin subunits, and the α_vβ₃ ligand, vitronectin, efficiently blocked the infectivity of the SA-dependent rhesus rotavirus RRV, its SA-independent variant nar3, and the neuraminidase-resistant human rotavirus strain Wa. Vitronectin and anti-β₃ antibodies, however, did not block the binding of virus to cells, indicating that rotaviruses interact with α_vβ₃ at a postbinding step, probably penetration. This interaction was shown to be independent of the tripeptide motif arginine-glycine-aspartic acid present in the natural ligands of this integrin. Transfection of CHO cells with α_vβ₃ genes significantly increased their permissiveness to all three rotavirus strains, and the increment of virus infectivity was reverted by incubation of these cells either with antibodies to β₃ or with vitronectin. These findings implicate α_vβ₃ integrin as a cellular receptor common to neuraminidase-sensitive and neuraminidase-resistant rotaviruses, and support the hypothesis that this integrin could determine, at least in part, the cellular susceptibility to rotaviruses.

Abstract

Rotaviruses, the leading cause of severe dehydrating diarrhea in infants and young children worldwide, are nonenveloped viruses that posses a genome of 11 segments of double-stranded RNA contained in a triple-layered protein capsid. The outermost layer is composed of two proteins, VP4 and VP7. VP4 forms spikes that extend from the surface of the virus, and it has been associated with a variety of functions, including initial attachment of the virus to the cell membrane and the penetration of the virion into the cell (1).

Rotaviruses have very specific cell tropism, infecting only enterocytes on the tip of intestinal villi (2), which suggests that specific host receptors must exist. In vitro, they also display restricted tropism, binding to a variety of cell lines, but efficiently infecting only those of renal or intestinal epithelium origin (3). Despite advances in knowledge regarding the molecular and structural biology of the virus, little is known about rotavirus cell receptors. It is known that some animal rotavirus strains attach to sialic acid (SA) on cell surfaces, and this interaction has been shown to be required for the efficient infection of virus to susceptible cells, both in vitro and in vivo (4). However, the binding of animal rotaviruses to an SA-containing cell receptor has been shown to be nonessential, because variants whose infectivity is no longer dependent on the binding to these acid sugars have been isolated (5). The secondary importance of SA as the attachment site for rotaviruses is also demonstrated by the fact that the infectivity of most, if not all, human rotavirus (HRV) strains is not affected by neuraminidase (NA) treatment of cells (6–8).

Integrins are a family of α/β heterodimers of cell adhesion receptors that mediate cell–extracellular matrix and cell–cell interactions, and are known to function as signaling receptors for a variety of cellular processes, including spreading, migration, proliferation, differentiation, and survival (9–11). These cell molecules are commonly used as receptors for many different viruses, including echoviruses 1, 8, 9, and 22 (12–15), coxsackievirus A9 (16), foot-and-mouth disease virus (17, 18), papillomavirus (19), adenovirus (20), adeno-associated virus type 2 (21), and hantaviruses (22), with integrin α_vβ₃ being, so far, the most frequently used as virus receptor (14, 16, 17, 20, 22).

Recently, it was found that rotavirus surface proteins contain sequence binding motifs for α₂β₁, α₄β₁, and α_xβ₂ integrins. Antibodies to these integrins, and peptides containing these sequence motifs, were shown to block the infectivity of simian rotavirus strain SA11 and the HRV strain RV5 (23). In addition, α₂β₁ and α₄β₁ integrins have been shown to mediate the attachment and entry of rotavirus SA11 into the human myelogenous leukemic cell line K562 (24).

We recently reported that proteins from MA104 cells, extracted with the nonionic detergent octyl β-glucoside under noncytolytic conditions, have the capacity to inhibit the infectivity of rotaviruses when preincubated with the virus before cell infection (25). In the present study, we have identified one of these proteins as the β₃ integrin subunit, and we demonstrate that α_vβ₃ integrin interacts with NA-sensitive and -resistant strains at a postattachment step and is capable of promoting rotavirus infection of the poorly permissive CHO (Chinese hamster ovary) cells.

Acknowledgments

We are grateful to Rafaela Espinosa and Pedro Romero for their excellent technical assistance, and to the computer staff, specially to Alma Martínez Valle, for their technical support. This work was partially supported by Grants 75197-527106 from the Howard Hughes Medical Institute, G0012-N9607 from the National Council for Science and Technology–Mexico, and IN201399 from the Dirección General de Apoyo al Personal Académico, Universidad Nacional Autónoma de México.

Acknowledgments

Abbreviations

SA	sialic acid
NA	neuraminidase
HRV	human rotavirus
RRV	rhesus rotavirus

Abbreviations

Footnotes

This paper was submitted directly (Track II) to the PNAS office.

Article published online before print: Proc. Natl. Acad. Sci. USA, 10.1073/pnas.250299897.

Article and publication date are at www.pnas.org/cgi/doi/10.1073/pnas.250299897

Footnotes

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