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Figure 1.8- Model for env mediated membrane fusion
An env trimer binds CD4. This results in conformational change in the gpl20 allowing the V3 loop to interact with the coreceptor. It is then proposed that this initiates conformational change in the entire env, including in gp4l which adopts a coiled-coil structure allowing fusion with the target cell. (Diagram adapted from Doms, R.W. and Peiper, S.C., Klasse, Virology (1997) 235, 179-190).
CCR5 that are important for this gpl 20 binding. These have involved using point mutations and chimeras between different chemokine receptors, both human and murine.
Murine CCR5 is 82% homologous to human CCR5 and can be activated by both human RANTES and M IP-1(3, but it is unable to function as a coreceptor for HIV-1. In the extracellular domains differences are found within the amino-terminus and first two loops, with the third loop being identical (Atchison et al, 1996, Bieniasz et al., 1997, Ross et at.,
1998). Analysis of murine/human chimeras showed that no single extracellular domain was sufficient to allow entry of all M-tropic viruses, but rather that all domains can play a role, and that which domain is important depends on the viral isolate used (Bieniasz et at.,
1997), although the amino-terminus and second loop appeared to be the most important ones (Atchison et al, 1996, Bieniasz et al, 1997, Ross et al, 1998). Unlike M-tropic virus, the dual-tropic viruses tested appeared to be very sensitive to any changes in any part of CCR5 (Bieniasz et al, 1997). The study of point mutations within murine CCR5 showed that relatively few amino acids need to be changed in order to render it functional (Ross et al, 1998). However, these functional mutations are spread out along all the extracellular region, further confirming that the interaction of HIV-1 and CCR5 is very complex and probably involves multiple contact sites with, and conformational change of, CCR5 (Ross et al, 1998).
Chimeras formed between CCR5 and other human chemokine receptors, such as CCR2b and CXCR4, have also been scrutinised (Doranz et al, 1997, Dragic et al, 1998, Lu et al, 1997, Rucker et al, 1996). These again indicate that all the extracellular domains may be involved, with the ones utilised varying between viral isolate used. These chimeras also showed that M-tropic virus is more forgiving of changes than dual-tropic virus, with these viruses being especially sensitive to changes and deletions within the amino-terminus (Doranz et al, 1997, Rucker et al, 1996).
1.5.2.4.2- Structure-function studies of CXCR4
As with CCR5, env and g p l20 can interact directly with CXCR4, independently of CD4. (Bandres et al, 1998). However again this interaction is considerably enhanced by the addition of sCD4, or deglycosylation of the g p l20 (Bandres et al, 1998). Indeed it appears that for either infection or membrane fusion and syncytium induction to take place CXCR4 alone is not sufficient, but that CD4 is required as well (Berson et al, 1996). The actual interaction of gpl 20 with CXCR4 appears not to be electrostatic as charged residues within the receptor play no role in HIV binding (Wang et al, 1998b).
Again the overall pattern of domain usage is similar to that of CCR5, with no one domain proving absolutely critical, and with different strains using slightly different areas (Brelot et al, 1997, Lu et al, 1997, Picard et al, 1997). Also the ability to signal and the presence of the cytoplasmic tail is not required (Lu et al, 1997). However some regions appear to be more important than others, as deletions within or mutations to the amino-
terminus, are well tolerated by almost all dual and T-tropic strains of HIV-1, although there are some exceptions (Picard et al, 1997). In addition, replacement of the third extracellular loop is tolerated similarly (Lu et al, 1997).
Of the remaining two domains, extracellular loops one and two, the second loop of CXCR4 seems to be the most important. Its replacement within CCR2 confers coreceptor ability on the hybrid receptor (Lu et al, 1997), and while rat CXCR4 can act as a coreceptor, the substitution of its second loop by the human equivalent allows a greater range of viruses to use it (Brelot et al, 1997). Thus the second loop seems to be of some importance for both dual and T-tropic strains of HIV-1. Nevertheless if Asp-187 within this loop is mutated to a neutral residue CXCR4 is able to support M-tropic infection (Wang et al, 1998b). This suggests that the viral interactions with the coreceptor are very subtle and probably based mainly on the conformation of the coreceptor, hence the fact that different strains interact predominantly with different loops.
1.5.2.5- Other chemokine receptors have been identified as coreceptors
In addition to the two major coreceptors for HIV-1, CCR5 and CXCR4 several other chemokine receptors have been identified that can act in a more limited capacity as coreceptors. These include CCR2b, CCR3, CCR9, the orphan receptors Bob, Bonzo, GPRl, Apj, and the CMV encoded chemokine receptor US28.
CCR2b has been shown to be used by some strains of HIV-1 and HIV-2 to gain entry in vitro (McKnight et al, 1998). A polymorphism has recently been found within CCR2b that may be protective against disease progression (Kostrikis et al, 1998), which is surprising since this coreceptor is rarely used in vitro. However not all reports agree that it has any protective effect (Eugen-Olsen et al, 1998).
CCR3 acts as a coreceptor for a subset of primary HIV-1 viruses that use CCR5, but is less efficient. CCR3 is sensitive to the chemokines eotaxin and RANTES, and infection can be inhibited by them in a dose-dependent manner (Choe et al, 1996). This receptor may of some importance in HIV-1 infection in the brain, being expressed at high levels on microglia (He et al, 1997). CCR9, on the other hand, has been shown to support the entry of only very few HIV-1 T-tropic viruses and so is of doubtful significance (Choe et al,
1998).
Of the orphan receptors. Bob and Bonzo appear to be the most important, mediating the entry of SIV as well as some M-tropic HIV-1 and HIV-2 strains (Alkhatib e ta l, 1997, Deng et a l, 1997, Liao et al, 1997). Apj, like CCR3 is found in relative abundance in the brain, but is also found on stimulated PBMC, however it can act as a coreceptor only for some dual and T-tropic strains of HIV-1, but not M-tropic stains that are commonly found in brain tissue, hence its significance is doubtful (Choe etal, 1998). Finally GPRl mediates the entry of SIV, but apparently is unable to mediate the entry of HIV-1 (Farzan et al,
The CMV-encoded CC chemokine receptor, US28, is functional both as a chemokine receptor, responding to the chemokines RANTES, M IP -la and MCP-1 (Gao & Murphy, 1994), and as an HIV-1 receptor, mediating both T- and M-tropic infection and cell-cell fusion (Pleskoff et al, 1997). It is also capable of enhancing cell-cell fusion mediated by other coreceptors such as CCR5, possibly by a mechanism distinct from that of its coreceptor function (Pleskoff e ta l, 1998). Given that CMV infections are a common finding among AIDS patients, and that CMV infects macrophages, the role of this receptor in the pathogenesis of HIV is of great interest.
All these coreceptors have been identified via in vitro methods, where cell surface expression levels are significantly higher than those found in vivo. Work has shown that cell surface levels of both the coreceptor and CD4 are of some significance as to whether a coreceptor may function effectively (Naif et al, 1998, Platt et al, 1998). Thus whether these minor coreceptors have an important role to play in vivo is unclear (Michael et al,
1998).