Polarized Human Immunodeficiency Virus Budding in Lymphocytes Involves a Tyrosine-Based Signal and Favors Cell-to-Cell Viral Transmission

Abstract

Maturation and release of human immunodeficiency virus type 1 (HIV-1) is targeted at the pseudopod of infected mononuclear cells. However, the intracellular mechanism or targeting signals leading to this polarized viral maturation are yet to be identified. We have recently demonstrated the presence of a functional YXXL motif for specific targeting of HIV-1 virions to the basolateral membrane surface in polarized epithelial Madin-Darby canine kidney cells (MDCK). Site-directed mutagenesis was used to demonstrate that the membrane-proximal tyrosine in the intracytoplasmic tail of the HIV-1 transmembrane glycoprotein (gp41) is an essential component of this signal. In the present study, immunolocalization of viral budding allowed us to establish that this tyrosine-based signal is involved in determining the exact site of viral release at the surface of infected mononuclear cells. Substitution of the critical tyrosine residue was also shown to increase the amount of envelope glycoprotein at the cell surface, supporting previous suggestions that the tyrosine-based motif can promote endocytosis. Although alteration of the dual polarization-endocytosis motif did not affect the infectivity of cell-free virus, it could play a key role in cell-to-cell viral transmission. Accordingly, chronically infected lymphocytes showed a reduced ability to transmit the mutant virus to a cocultivated cell line. Overall, our data indicate that the YXXL targeting motif of HIV is active in various cell types and could play an important role in viral propagation; this may constitute an alternative target for HIV therapeutics and vaccine development.

The C-terminal tail of the gp41 transmembrane envelope glycoprotein of HIV-1 clades A, B, C, and D may exist in two conformations: an analysis of sequence, structure, and function

“…However, we shall propose that only the minor 3-MSD form is incorporated into virions. These proposals are consistent with, and rationalize the observed degradation of the majority (85 -95%) of the1-MSD form of cellular gp160 (Bultmann et al, 2000;Courageot et al, 1999;Jabbar and Nayak, 1990;Pfeiffer et al, 1997;Willey et al, 1988), the apparently contradictory evidence of a functioning first tyrosine-sorting signal (Berlioz-Torrent et al, 1999;Boge et al, 1998;Deschambeault et al, 1999;Egan et al, 1996;Ohno et al, 1997;Rowell et al, 1995;West et al, 2002) and basolateral-sorting signal (Deschambeault et al, 1999;Lodge et al, 1997;Owens et al, 1991), with the immunogenic and antigenic properties of the Kennedy region (Chanh et al, 1986;Dalgleish et al, 1988;Evans et al, 1989;Ho et al, 1987;Kennedy et al, 1986;Niedrig et al, 1992).…”

“…Thus, the first tyrosine-sorting signal ( 719 YSPL 722 ) could interact with the AP-1 and AP-3 complexes in the TGN and target gp41 to the lysosomes. However, as stated above, this signal functions at the plasma membrane (Berlioz-Torrent et al, 1999;Boge et al, 1998;Deschambeault et al, 1999;Egan et al, 1996;Ohno et al, 1997;Rowell et al, 1995;West et al, 2002), and is not known to be active in the TGN. It may be that the amount of gp41 synthesized saturates the lysosomal targeting system in the TGN, allowing gp41 to reach the cell surface and the first tyrosine-sorting signal to function as an endocytosis signal.…”

“…The definition and values of the peaks obtained in the above analysis make the data highly significant. The position and number of residues in the MSDs and other regions of interest are summarized in Table 1 and 1999; Egan et al, 1996;Ohno et al, 1997;Rowell et al, 1995;West et al, 2002) and basolateral sorting (Deschambeault et al, 1999;Lodge et al, 1997;Owens et al, 1991).…”

“…Also for signal functionality there is a strict requirement that the tyrosine residue is the 7th -11th residue from the membrane (Rohrer et al, 1996). Gp41 is also involved in basolateral localization of envelope protein in the plasma membrane of polarized cells (Owens et al, 1991), a property that is lost when the tyrosine of the first signal is substituted (Deschambeault et al, 1999;Lodge et al, 1997). In the 1-MSD model of gp41, Gallaher et al (1989) have proposed that the MSD ends at residue 712 V, and thus 719 Y will be the 7th residue from the membrane, and Fig.…”

“…The amount of gp160 secreted from the cell is less than 10% of the gp120 secreted (Willey et al, 1988), although the amount varies with the type of host cell (Moulard et al, 1999). Tyrosine-dependent sorting signals (YxxB, where x represents any amino acid residue and B represents a bulky hydrophobic residue), and possibly di-leucine-sorting signals are found in the C-terminal tail of gp41, and have been implicated in transport of gp160 and gp120-gp41 (see Results and discussion section and Berlioz-Torrent et al, 1999;Boge et al, 1998;Bu et al, 2004;Deschambeault et al, 1999;Egan et al, 1996;Lodge et al, 1997;Ohno et al, 1997;Owens et al, 1991;Rowell et al, 1995;West et al, 2002;Wyss et al, 2001).…”