Kaposi’s sarcoma-associated herpesvirus glycoprotein K8.1 is critical for infection in a cell-specific manner and functions at the attachment step on keratinocytes

Abstract

Kaposi's sarcoma-associated herpesvirus (KSHV) is associated with Kaposi's sarcoma and several B cell malignancies. K8.1, the major antigenic component of the KSHV virion, has been reported to play a critical role in the infection of certain B cells, but otherwise its function remains enigmatic. We created a K8.1 knockout virus (KSHV∆K8.1) in the BAC16 genetic background and analyzed its infectivity on a range of adherent cells. We observed a strong defect on several epithelial cells, e.g. the HaCaT keratinocyte model cell line, HEK 293T and A549 lung epithelial cells, but no such defect on other cells, among them e.g. lymphatic and blood endothelial cells. Mechanistically, we found that reduced infectivity of the K8.1 knockout virus correlated with reduced attachment to HaCaT cells. The defect in infectivity of KSHV∆K8.1 could be rescued by complementation through expression of K8.1 in KSHV∆K8.1 producing cells by means of a lentiviral vector. In a coculture infection model, KSHV∆K8.1 was highly efficient at infecting the BJAB B cell line but was significantly impaired at infecting the MC116 B cell line, in line with a previous report. In fusion assays together with the gH/gL glycoprotein complex and gB, the components of the conserved herpesviral core fusion machinery, we did not observe activation of membrane fusion by K8.1 or its R8.1 homolog of the rhesus monkey rhadinovirus. In summary, we found K8.1 to function in a highly cell-specific manner during KSHV entry at the attachment step, playing an important role in the infection of epithelial cells.

Tim-1 and Tim-4 mediate entry of the human Kaposi’s sarcoma-associated herpesvirus and the related rhesus monkey rhadinovirus

“…The attachment assay was performed as described previously 25 . Briefly, cells were incubated with KSHV and spin-inoculated by centrifugation at 4°C and 4122g for 30min.…”

“…These interactions are generally thought to result in endocytosis of the viral particle and then membrane fusion from endocytic compartments, as various endocytic routes have been found to be important for productive entry of KSHV and it is sensitive to inhibition of the vacuolar ATPase 14,16–18 . For both KSHV and the related rhesus monkey rhadinovirus (RRV) experiments aimed at ablating certain receptor interactions, such as those with integrins 16,19 or Eph family receptors 11,14,20–22 , mutating receptor interaction sites 12,21 or deleting individual glycoproteins 23–25 did not result in complete loss of infectivity, which suggests the existence of additional cellular factors that mediate entry. KSHV and RRV exhibit high similarity with regard to cell tropism, associated disease spectrum and in part receptor usage as both viruses use Eph family receptors 12,14,22 .…”