The T-Cell Receptor ζ Chain Contains Two Homologous Domains with Which Simian Immunodeficiency Virus Nef Interacts and Mediates Down-Modulation

Abstract

We have recently demonstrated that simian immunodeficiency virus (SIV) Nef binds to the ζ chain of the T-cell receptor (TCR), leading to its down-modulation from T-cell surfaces (I. Bell, C. Ashman, J. Maughan, E. Hooker, F. Cook, and T. A. Reinhart, J. Gen. Virol. 79:2717–2727, 1998). Using a panel of human as well as rhesus macaque TCR ζ cytoplasmic domain mutants, we have identified in this report two linear peptides in the cytoplasmic domain of TCR ζ which independently interact with SIV Nef. Each SIV Nef interaction domain was sufficient in the absence of the other for interaction with SIV Nef in a yeast two-hybrid assay. In parallel, we demonstrated that Nef down-modulation of CD8-TCR ζ fusion proteins containing full-length or truncated portions of the TCR ζ cytoplasmic domain occurs in transiently transfected 293T cells. Furthermore, using proteins expressed inEscherichia coli, a glutathioneS-transferase–Nef fusion protein coprecipitated histidine-tagged portions of the TCR ζ cytoplasmic domain which contained SNID-1 or SNID-2. The peptides targeted by SIV Nef, YNELNL and YSEIGMKGERRR, are portions of the first and second of three immunoreceptor tyrosine-based activation motifs which are important in signal transduction, thymocyte development, and TCR biogenesis. These results demonstrate that SIV Nef binds to two distinct domains on TCR ζ in the absence of other T-cell-specific factors, and that interaction with either domain is sufficient to cause down-modulation of TCR ζ.