HIV-1 Nef AND HOST CELL PROTEIN KINASES

Kalle Saksela

Institute of Medical Technology, University of Tampere, P.O. Box 607, FIN-33101, Tampere, Finland

Received 11/10/97 Accepted 11/17/97

4. CONCLUDING REMARKS

Despite the recent advances in research on Nef, its critical cellular function and molecular mechanism of action in the pathogenesis of AIDS remain unclear. A number of observations suggest that physical and functional interactions with host cell protein kinases are important for Nef function. However, the relevant protein kinases in various target cells of HIV infection still await for definitive identification, posing a major challenge for future investigations.

Molecular cloning of the Nef-associated serine kinase activity, NAK, would seem necessary in order to conclusively address its possible involvement in the cell biology of Nef. The role(s) of different isoforms of PKC as effectors of Nef will also need to be carefully elucidated. On the other hand, the potential role and the downstream molecular events of the Nef/Hck interaction in providing HIV-infected macrophages with a capacity to promote HIV infection alsio requires furtehr experimental attention. Also, the suggestive but contradictory reports on an interaction of Nef with the T cell-specific Src-kinase Lck will have to be followed-up in more detail. If indeed both Hck and Lck turn out to be important, it will be interesting to see if they serve corresponding functions, or whether Nef has evolved to employ them for different purposes in these two cell types.

Although the number of suggested Nef-interacting host cell proteins has rapidly increased, it is likely that not all cellular partners of Nef have yet been identified. In this regard, search for novel high-affinity Nef-binding SH3 domain-containing proteins in non-myeloid cells, such as T lymphocytes, poses one attractive area for further investigations. Besides host proteins involved in signal transduction, other types of novel Nef-interacting proteins are also likely to be discovered, as all pieces in the puzzle regarding the Nef-induced downregulation of CD4 and MHC I are probably not yet at hand.

While the identification of the relevant Nef-binding host cell proteins will undoubtedly accelerate the discovery the critical cellular processes which Nef modulates in order to mediate its pathogenic function, the reverse argument can also be made. A better idea of the germane Nef-induced changes in cellular physiology would greatly help to decide which ones of the candidate Nef-binding host cell proteins are in fact relevant. In this regard, our current level of understanding leaves a lot of room for improvement. For example, it is still unclear if the true role of Nef is to inhibit or enhance the intracellular signaling mediated through the T cell receptor complex, or whether depending on circumstances both might occur, or even, if both of these effects merely represent different artifacts of ectopic Nef overexpression. Given the likely role of Nef as a regulator of cellular signaling pathways, it is possible that the common practice of using established cell lines to study the effects of Nef on cellular physiology may not have been optimal, and study systems which would better mimick Nef expression during natural HIV infection may have to be considered more carefully in the future.

SIV infection of rhesus macaques has been regarded as the ultimate test for the functionality of various Nef alleles, and it is indeed the model in which the requirement for Nef in the pathogenesis of AIDS was first demonstrated. In this model, unlike in cell culture systems or in SCID-hu mice, the interplay between the virus and the immune system can also be assessed which (besides being important for any infectious disease) might be relevant to appropriately reveal the in vivo function of Nef. The possibility to examine the reversion of mutations in Nef which inhibit the replicative potential of the virus provides another valuable experimental tool in this model. However, besides the broader question of the overall similarity of simian AIDS in SIV-infected macaques compared to AIDS in HIV-infected humans (target cell populations, immunopathogenesis, etc.), there are indications, as discussed in the previous chapter, suggesting that SIV Nef might differ from HIV-1 Nef in certain functional aspects. Such differences, even if subtle, could significantly complicate the use of this model for testing of hypotheses based on studying HIV-1 Nef.

The lack of complete functional correspondence of HIV-1 and SIV Nef proteins is also suggested by the efforts to create a pathogenic SIV strain in which the Nef gene has been transplanted from HIV-1. While such exercises involving exchanges of much larger fragments from other regions of these genomes have been quite successful, substitution of SIV Nef with HIV-1 Nef has proved difficult, and positive results on this have been obtained only recently (P. Luciw, personal communication). The development of such chimeric SHIV viruses represents a significant advance, and should help to reduce the possible problems arising from dissimilarity between HIV-1 and SIV Nef proteins. It should be noted, however, that accumulation of approximately half a dozen amino acid changes in certain positions of the transplanted HIV-1 Nef are required for adaptation of these SHIVs to become pathogenic in macaques.

Besides providing an improved model for in vivo testing of the functional importance of specific mutations introduced into the HIV-1 Nef, such SHIV viruses might prove valuable in evaluating possible anti-HIV therapeutic agents targeted against Nef. It is probably realistic to deem that in order to ensure continuous agreeable progress in antiretroviral therapy against AIDS, new drugs against the currently targeted as well as other HIV proteins need to be developed. The demonstrated role of Nef in the pathogenesis of AIDS, together with the improved understanding about the molecular details of HIV-1 Nef structure and function make Nef an attractive target for anti-HIV drug development. The necessity for HIV-1 Nef protein, in order to remain functional, to preserve its architecture in critical regions which interact with host cell proteins, such as the Nef SH3-binding surface, present feasible submolecular targets within the Nef protein, and suggest that the therapeutic effects of drugs aimed at such sites might be less prone to be overcome by mutated drug resistant viruses than is the case with the current antiretroviral therapies.