Anna Segerman

Adenovirus Type 11 Uses CD46 as a Cellular Receptor

ABSTRACT The 51 human adenovirus serotypes are divided into six species (A to F). Many adenoviruses use the coxsackie-adenovirus receptor (CAR) for attachment to host cells in vitro. Species B adenoviruses do not compete with CAR-binding serotypes for binding to host cells, and it has been suggested that species B adenoviruses use a receptor other than CAR. Species B adenoviruses mainly cause disease in the respiratory tract, the eyes, and in the urinary tract. Here we demonstrate that adenovirus type 11 (Ad11; of species B) binds to Chinese hamster ovary (CHO) cells transfected with CD46 (membrane cofactor protein)-cDNA at least 10 times more strongly than to CHO cells transfected with cDNAs encoding CAR or CD55 (decay accelerating factor). Nonpermissive CHO cells were rendered permissive to Ad11 infection upon transfection with CD46-cDNA. Soluble Ad11 fiber knob but not Ad7 or Ad5 knob inhibited binding of Ad11 virions to CD46-transfected cells, and anti-CD46 antibodies inhibited both binding of and infection by Ad11. From these results we conclude that CD46 is a cellular receptor for Ad11.

Adenovirus Types 11p and 35p Show High Binding Efficiencies for Committed Hematopoietic Cell Lines and Are Infective to These Cell Lines

ABSTRACT Hematopoietic cells are attractive targets for gene therapy. However, no satisfactory vectors are currently available. A major problem with the most commonly used adenovirus vectors, based on adenovirus type 2 (Ad2) or Ad5, is their low binding efficiency for hematopoietic cells. In this study we identify two adenovirus serotypes with high affinity for hematopoietic cells. The binding efficiency of prototype serotypes Ad4p, Ad11p, and Ad35p for different committed hematopoietic cell lines representing T cells (Jurkat), B cells (DG75), monocytes (U937-2), myeloblasts (K562), and granulocytes (HL-60) was evaluated and compared to that of Ad5v, the commonly used adenovirus vector, using flow cytometry. In contrast to Ad5v, which bound to less than 10% of the cells in all experiments, Ad11p and Ad35p showed high binding efficiency for all of the different hematopoietic cell lines. Ad4p bound to the lymphocytic cell lines to some extent but less well to the myelomonocytic cell lines. The abilities of the different serotypes to infect, replicate, and form complete infectious particles in the hematopoietic cell lines were also investigated by immunostaining, 35S labeling of viral proteins, and titrations of cell lysates. Ad11p and Ad35p infected the highest proportion of cells, and Ad11p infected all of the cell lines investigated. The Ad11p hexon was expressed equally well in K562 and A549 cells. Jurkat cells also showed high levels of expression of Ad11p hexons, but the production of infectious particles was low. The binding properties of virions were correlated to their ability to infect and be expressed.

There Are Two Different Species B Adenovirus Receptors: sBAR, Common to Species B1 and B2 Adenoviruses, and sB2AR, Exclusively Used by Species B2 Adenoviruses

ABSTRACT Unlike most adenovirus (Ad) serotypes, the species B Ads do not use the coxsackie-adenovirus receptor as an attachment receptor. The species B attachment receptor(s) has not yet been identified and is also poorly characterized. Species B Ads can be further divided into species B1 and B2 Ads, and these display different organ tropisms, suggesting a difference in receptor usage. We have studied the receptor interactions of the species B1 serotypes 3p and 7p and the species B2 serotypes 11p and 35 and characterized the properties of the species B receptor(s). Reciprocal blocking experiments using unlabeled Ad11p or Ad3p virions to block the binding to A549 cells of 35S-labeled 3p, 7p, 11p, and 35 showed that only Ad11p virions efficiently blocked the binding of all the species B Ads studied (≥70%). Thus, there is apparently a common species B Ad receptor (sBAR). However, Ad3p virions only partially (≤30%) blocked the binding of Ad11p and Ad35 to A549 cells. Binding experiments after trypsin treatment of the cells confirmed that the species B2 serotypes address at least two different receptors on A549 and J82 cells, since sBAR is trypsin sensitive but the species B2 Ad receptor (sB2AR) is not. Both receptors are proteins or glycoproteins, since binding of all species B serotypes was abolished after proteinase K or subtilisin treatment of A549 or J82 cells. Furthermore, binding of the species B serotypes to sBAR was abolished with EDTA and restored with Ca2+, whereas the binding of Ad11p and Ad35 to SB2AR was independent of divalent cations.

The Arg279Gln Substitution in the Adenovirus Type 11p (Ad11p) Fiber Knob Abolishes EDTA-Resistant Binding to A549 and CHO-CD46 Cells, Converting the Phenotype to That of Ad7p.

ABSTRACT The major determinant of adenovirus (Ad) attachment to host cells is the C-terminal knob domain of the trimeric fiber protein. Ad type 11p (Ad11p; species B2) in contrast to Ad7p (species B1) utilizes at least two different cellular attachment receptors, designated sBAR (species B adenovirus receptor) and sB2AR (species B2 adenovirus receptor). CD46 has recently been identified as one of the Ad11p attachment receptors. However, CD46 did not seem to constitute a functional receptor for Ad7p. Although Ad7p shares high knob amino acid identity with Ad11p, Ad7p is deficient in binding to both sB2AR and CD46. To determine what regions of the Ad11p fiber knob are necessary for sB2AR-CD46 interaction, we constructed recombinant fiber knobs (rFK) with Ad11p/Ad7p chimeras and Ad11p sequences having a single amino acid substitution from Ad7p. Binding of the constructs to A549 and CHO-CD46 BC1 isoform-expressing cells was analyzed by flow cytometry. Our results indicate that an Arg279Glu substitution is sufficient to convert the Ad11p receptor-interaction phenotype to that of Ad7p and abolish sB2AR and CD46 interaction. Also a Glu279Arg substitution in Ad7p rFKs increases CD46 binding. Thus, the lateral HI loop of the Ad11p fiber knob seems to be the key determinant for Ad11p sB2AR-CD46 interaction. This result is comparable to another non-coxsackie-adenovirus receptor binding Ad (Ad37p), where substitution of one amino acid abolishes virus-cell interaction. In conjunction with previous results, our findings also strongly suggest that sB2AR is equivalent to CD46.