Human adenovirus 52 (HAdV-52) is one of only three known HAdVs equipped with both a long and a short fiber protein. While the long fiber binds to the coxsackie and adenovirus receptor, the function of the short fiber in the virus life cycle is poorly understood. Here, we show, by glycan microarray analysis and cellular studies, that the short fiber knob (SFK) of HAdV-52 recognizes long chains of α-2,8-linked polysialic acid (polySia), a large posttranslational modification of selected carrier proteins, and that HAdV-52 can use polySia as a receptor on target cells. X-ray crystallography, NMR, molecular dynamics simulation, and structure-guided mutagenesis of the SFK reveal that the nonreducing, terminal sialic acid of polySia engages the protein with direct contacts, and that specificity for polySia is achieved through subtle, transient electrostatic interactions with additional sialic acid residues. In this study, we present a previously unrecognized role for polySia as a cellular receptor for a human viral pathogen. Our detailed analysis of the determinants of specificity for this interaction has general implications for protein–carbohydrate interactions, particularly concerning highly charged glycan structures, and provides interesting dimensions on the biology and evolution of members of Human mastadenovirus G. Significance We present here that adenovirus type 52 (HAdV-52) attaches to target cells through a mechanism not previously observed in other human pathogenic viruses. The interaction involves unusual, transient, electrostatic interactions between the short fiber capsid protein and polysialic acid (polySia)-containing receptors on target cells. Knowledge about the binding interactions between polySia and its natural ligands is relatively limited, and our results therefore provide additional insight not only into adenovirus biology but also into the structural basis of polySia function. Since polySia can be found in high expression levels in brain and lung cancers where its presence is associated with poor prognosis, we suggest that this polySia-binding adenovirus could be useful for design of vectors for gene therapy of these cancers.

Significance: The adenovirus capsid protein is built by three main capsomers: hexon, fiber, and penton base. Entry is mediated by fiber proteins anchoring the virus to host cell receptors and is followed by penton base proteins engaging coreceptors, resulting in entry. Here, we demonstrate that human adenovirus species D types, which constitute two-thirds of all human adenoviruses, enter host cells through a direct interaction between the hexon protein and CD46. This study provides insights into the entry mechanisms used by human adenoviruses. As these viruses are also used as vaccine vectors for prevention of other infectious diseases, the results provided will also be useful for further development of human adenovirus species D types as vaccine vectors.

Significance: Acute hemorrhagic conjunctivitis (AHC) is a painful and highly contagious infection of the eye, with reported incidence rates of up to 48%. No drugs or vaccines are available for treatment or prevention of AHC. Coxsackievirus A24 variant (CV-A24v) is the main etiological agent of AHC, being responsible for >10 million AHC cases worldwide during the last decades. We have identified the CV-A24v protein receptor and determined the high-resolution structure of the virus–receptor complex. Furthermore, we found that an adaptation which enhances binding to the receptor sialic acid may have contributed to the pathogenicity and pandemic nature of CV-A24v. These findings highlight the importance of sialic acid for viruses with ocular tropism, such as influenza A virus and several adenoviruses.

A cryo-EM structure reveals unique traits of a human adenovirus that has acquired gastrointestinal tropism.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by rich deposits of extracellular matrix (ECM), affecting the pathophysiology of the disease. Here, we identified galectin 4 (gal 4) as a cancer cell–produced protein that was deposited into the ECM of PDAC tumors and detected high-circulating levels of gal 4 in patients with PDAC. In orthotopic transplantation experiments, we observed increased infiltration of T cells and prolonged survival in immunocompetent mice transplanted with cancer cells with reduced expression of gal 4. Increased survival was not observed in immunodeficient RAG1−/− mice, demonstrating that the effect was mediated by the adaptive immune system. By performing single-cell RNA-sequencing, we found that the myeloid compartment and cancer-associated fibroblast (CAF) subtypes were altered in the transplanted tumors. Reduced gal 4 expression associated with a higher proportion of myofibroblastic CAFs and reduced numbers of inflammatory CAFs. We also found higher proportions of M1 macrophages, T cells, and antigen-presenting dendritic cells in tumors with reduced gal 4 expression. Using a coculture system, we observed that extracellular gal 4 induced apoptosis in T cells by binding N-glycosylation residues on CD3ε/δ. Hence, we show that gal 4 is involved in immune evasion and identify gal 4 as a promising drug target for overcoming immunosuppression in PDAC.

The enteric species F human adenovirus types 40 and 41 (HAdV-40 and -41) are the third most common cause of infantile gastroenteritis in the world. Knowledge about HAdV-40 and -41 cellular infection is assumed to be fundamentally different from that of other HAdVs since HAdV-40 and -41 penton bases lack the αV-integrin-interacting RGD motif. This motif is used by other HAdVs mainly for internalization and endosomal escape. We hypothesised that the penton bases of HAdV-40 and -41 interact with integrins independently of the RGD motif. HAdV-41 transduction of a library of rodent cells expressing specific human integrin subunits pointed to the use of laminin-binding α2-, α3- and α6-containing integrins as well as other integrins as candidate co-receptors. Specific laminins prevented internalisation and infection, and recombinant, soluble HAdV-41 penton base proteins prevented infection of human intestinal HT-29 cells. Surface plasmon resonance analysis demonstrated that HAdV-40 and -41 penton base proteins bind to α6-containing integrins with an affinity similar to that of previously characterised penton base:integrin interactions. With these results, we propose that laminin-binding integrins are co-receptors for HAdV-40 and -41.