Human immunodeficiency virus infection is efficiently mediated by a glycolipid-anchored form of CD4.

Abstract

Two broad roles have been revealed for the CD4 molecule. It serves as a receptor for both class II major histocompatibility complex molecules and human immunodeficiency virus (HIV). Upon binding class II major histocompatibility molecules, CD4 functions to enhance T-cell activation. By binding to CD4, HIV gains entry into the cell. We have used a chimeric molecule of CD4 and lymphocyte functionassociated antigen 3 (LFA-3), CD4PI, which lacks a membrane-spanning domain and is instead anchored in the membrane by linkage to glycosyl-phosphatidylinositol. (10)(11)(12), and perhaps induce inhibitory signals (12). Previous work has shown that alteration or replacement of various cytoplasmic and membrane-spanning regions of CD4, as well as deletions of the cytoplasmic domain, can disrupt modulation by phorbol 12-myristate 13-acetate (PMA) (13-15) but not the inhibitory effects ofgpl20 (10) or the ability to mediate infection by HIV (13,14). While these mutants show that the wild-type CD4 sequence of these regions is not required by HIV, they do all have membrane-spanning and, at least, rudimentary cytoplasmic domains, thus leaving open the possibility of a generic requirement for such structures in order to serve as a viral receptor. The retention of membrane-spanning sequences may allow continued interactions with other membrane-spanning regions. Likewise, even the few cytoplasmic amino acids that are retained may allow interactions with cytoskeletal or other proteins at the inner surface of the membrane. Any of these interactions could be important for the uptake of virus, which must follow binding.Here we demonstrate the ability of CD4PI, a lipidanchored form of CD4, to mediate productive infection by HIV, thereby proving that neither the membrane-spanning nor the cytoplasmic domain of CD4 is required, specifically or as a structural element, in order for CD4 to function as a viral receptor. These results serve to identify the class of naturally glycosyl-phosphatidylinositol (GPI)-anchored proteins as potential viral receptors. MATERIALS AND METHODSMedia. All experiments were carried out in RPMI 1640 supplemented with 10% calf serum, 10 mM Hepes, 2 mM L-glutamine, 50 ,RM 2-mercaptoethanol, 100 units ofpenicillin per ml, and 100 Ag of streptomycin per ml except as noted otherwise. Once established, the HSB2-derived lines were carried in the same medium but with only 5% serum.Expression of CD4 and CD4PI in HSB-2 Cells. Expression of the CD4 constructs in the human T-cell line HSB-2 was obtained by coculture with the retroviral producer lines MNST4 DAMP (6) and MNCT4PI DAMP as described (6,15,16). In brief, the retroviral producer lines were generated by transfection, by calcium phosphate coprecipitation, of the amphotropic helper line DAMP with defective proviral vector DNA carrying neomycin resistance and the cDNA encoding CD4 or CD4PI, followed by selection with the antibiotic G418 (1 mg/ml). Following coculture the HSB-2 infectants were separated from the adherent producer line by repeated plating and subj...

The V domain of herpesvirus Ig-like receptor (HIgR) contains a major functional region in herpes simplex virus-1 entry into cells and interacts physically with the viral glycoprotein D

“…Thus, HIgR adds a new member to the list of Ig-like viral receptors whose major functional domains reside in the more external regions. The list includes PVR, the poliovirus receptor, ICAM, the rhinovirus receptor, and CD4, the HIV receptor (29)(30)(31)(32)(33)(34)(35)(36). That the N-terminal domains carry functional regions is consistent with the view that, even in the folded molecules, these regions are located more externally and therefore are more likely to interact with the appropriate regions of the virions.…”

Specific cell surface requirements for the infection of CD4-positive cells by human immunodeficiency virus types 1 and 2 and by simian immunodeficiency virus

“…(1) Mouse cells of different lineage expressing human CD4 bind virus but infection is blocked before fusion (Maddon et al, 1986) . CD4 constructs with truncated cytoplasmic sequences, with substituted murine CD4 cytoplasmic domains, or with the two CD4 N-terminal domains linked to the cell membrane by a glycophosphotidyl inositol tail confer susceptibility to human HeLa cells but not to mouse cells (Bedinger et al ., 1988 ;Maddon et al-, 1988 ;Diamond et al ., 1990) .…”

Intercellular transfer of a glycosylphosphatidylinositol (GPI)-linked protein: release and uptake of CD4-GPI from recombinant adeno-associated virus-transduced HeLa cells.

“…HIV efficiently infects human cells having endogenously expressed GPI-anchored CD4 receptors (8,9 (11).…”