Irreversible eclipse of poliovirus by HeLa cells

Abstract

The binding of poliovirus to receptors of either intact or disrupted human cells results in an apparent eclipse of infectivity immediately upon adsorption. However, this is not a true eclipse, for full virus infectivity is restored by treatment with a number of agents (by very concentrated salt solutions, or by hydrogen bond-disrupting agents such as 8 M urea, or by very low pH). Of those agents capable of reversing the binding between virus and cell, only concentrated salt solutions (e.g., 6 M LiCl) did not also destroy the virus-binding ability of receptor protein, a result supporting the concept that electrostatic forces may be the major binding forces in specific adsorption of poliovirus and other enteroviruses. No evidence for disulfide bridging between cell and virus in adsorption or penetration was obtained using sulfhydryl compounds. Trypsin digestion, which readily destroyed poliovirus receptor activity of HeLa cell microsomes nevertheless failed to release virus already bound to receptor, indicating that adsorbed virus is in close proximity to the trypsin-sensitive receptor protein. Poliovirus attached to living HeLa cells, when tested at intervals for restoration of infectivity with 6 M LiCl or 8 M urea, lost infectivity exponentially upon incubation at 37° but not at 0°. This appears to be a “true eclipse,” and it did not occur following adsorption of virus to receptors of disrupted cells. This eclipse of virus parallels virus “penetration” as measured by loss of antiserum sensitivity of adsorbed virus at intervals. At no time was most of the cell-bound virus found to have penetrated beyond the cell surface (beyond antiserum sensitivity) and to have retained infectivity recoverable with salt or urea treatment. This finding suggested that poliovirus is eclipsed either at the cell surface or very soon after having left the surface. Attempts to block virus eclipse with metabolic inhibitors were not successful. A poliovirus-resistant HeLa cell population failed to eclipse virus at the normal rate, and most uneclipsed virus remained at the cell surface. Even disrupted HeLa cells were able to eclipse poliovirus infectivity, but only when the cells were disrupted after virus attachment to receptors. Finally, it was found that cell membrane fractions of HeLa cell homogenates (membranes which sediment at low speeds) adsorbed and eclipsed poliovirus, whereas microsomal membranes adsorbed, but failed to eclipse poliovirus. It is concluded that irreversible eclipse of poliovirus may occur at the cell surface following reversible attachment to cell receptors, and that one of the consequences of this eclipse may be loss of antiserum sensitivity of the virus-cell complex.