Understanding internalization of rotavirus VP6 nanotubes by cells: towards a recombinant vaccine

Abstract

Rotavirus VP6 nanotubes are an attractive option for a recombinant vaccine against rotavirus disease. Protection against rotavirus infection and an adjuvant effect have been observed upon immunization with VP6 nanotubes. However, little information exists on how VP6 nanotubes interact with cells and trigger an immune response. In this work, the interaction between VP6 nanotubes and different cell lines was characterized. VP6 nanotubes were not cytotoxic to any of the animal or human cell lines tested. Uptake of nanotubes into cells was cell-line-dependent, as only THP1 and J774 macrophage cells internalized them. Moreover, the size and spatial arrangement of VP6 assembled into nanotubes allowed their uptake by macrophages, as double-layered rotavirus-like particles also displaying VP6 in their surface were not taken up. The internalization of VP6 nanotubes was inhibited by methyl-β-cyclodextrin, but not by genistein, indicating that nanotube entry is specific, depends on the presence of cholesterol in the plasma membrane, and does not require the activity of tyrosine kinases. The information generated here expands our understanding of the interaction of protein nanotubes with cells, which is useful for the application of VP6 nanotubes as a vaccine.

Rotavirus VP6 preparations as a non-replicating vaccine candidates

“…Different assemblies of VP6 have been defined based on pH and protonation state that spherical particles were formed in the pH range 3.5 ± 5.5, large tubes in the range 5.5 ± 7, and small tubes above pH 7 [29]. These VP6 structures are able to protect against RV-induced diarrhea [149][150][151][152] although VP6 trimers elicited low level of Ab response in immunized mice [150]. In particular, elongated tubular structures of VP6 not only induce higher level of immunogenicity compared with trimers or spherical structures of VP6 [149,151], but also may have the potential to act as an adjuvant [152,153] suggesting the reliance of the Ab response induction on the protein quaternary structure [154].…”

“…These VP6 structures are able to protect against RV-induced diarrhea [149][150][151][152] although VP6 trimers elicited low level of Ab response in immunized mice [150]. In particular, elongated tubular structures of VP6 not only induce higher level of immunogenicity compared with trimers or spherical structures of VP6 [149,151], but also may have the potential to act as an adjuvant [152,153] suggesting the reliance of the Ab response induction on the protein quaternary structure [154]. To support this, a comparison of the immunogenicity of recombinant tubular VP6 and DLP2/6 [24] has shown that both tubular VP6 and DLP2/6 are able to induce similar effective humoral and cellular responses against RV in mice [24].…”

“…At least eight groups or species (A to H) have been differentiated [28]. VP6 expressed alone is capable of self-assembling into VLPs or tubules termed nanotubules and trimers [29][30][31]. It has also been shown that nanotubules can induce strong immune response [32].…”

Rotavirus capsid VP6 tubular and spherical nanostructures act as local adjuvants when co-delivered with norovirus VLPs

“…VP6 nanotubes (VP6T) and nanospheres (VP6S), when expressed in vitro [12,[24][25][26][27] under different conditions [25,28]. The rVP6 is highly immunogenic [9,12,29], and it has also been used as a carrier or delivery platform for heterologous protein antigens and genetically fused epitope-based vaccines, with improved response to the foreign antigen [30][31][32][33][34]. VP6 is stable at different pH conditions, and when delivered orally it was targeted to intestinal cells, offering a promising new delivery platform to transport pharmaceutical compounds to gastrointestinal tract [35].…”

Rotavirus VP6 as a potential vaccine candidate

“…Co‐administration of VP6 nanotubes with norovirus VLPs not only induced protective immunity against RV infection but also showed an adjuvant effect of immune response for norovirus VLPs . To support this, Rodriguez et al has investigated the adjuvant effect of VP6 nanotubes and showed that nanotubes are taken up efficiently by macrophages and dendritic cells in vitro, inducing several proinflammatory cytokines and chemokines . VP6 nanotubes were also indicated to support the internalization of co‐delivered norovirus VLPs to the antigen presenting cells .…”