CHARACTERIZATION OF THE NUCLEAR EXPORT SIGNAL OF HUMAN T-CELL LYMPHOTROPIC VIRUS TYPE-1 REX REVEALS THAT NUCLEAR EXPORT IS MEDIATED BY POSITION-VARIABLE HYDROPHOBIC INTERACTIONS

We previously determined that amino acids 64 to 120 of human T-cell ly mphotropic virus type 1 (HTLV-1) Rex can restore the function of an ef fector domain mutant of human immunodeficiency virus type 1 (HIV-1) Re v (T. J. Hope, B. L. Bond, D. McDonald, N. P. Klein, and T. G. Parslow , J. Virol. 65:6001-6007, 1991). In this report, we (i) identify and c haracterize a position-independent 17-amino-acid region of HTLV-1 Rex that fully complements HIV-1 Rev effector domain mutants and (ii) show that this 17-amino-acid region and specific hydrophobic substitutions can serve as nuclear export signals. Mutagenesis studies revealed tha t four leucines within the minimal region were essential for function. Alignment of the minimal Rex region with the HIV-1 Rev effector domai n suggested that the position of some of the conserved leucines is fle xible. We found two of the leucines could each occupy one of two posit ions within the context of the full-length HTLV-1 Rex protein and main tain function. The idea of flexibility within the Rex effector domain was confirmed and extended by identifying functional substitutions by screening a library of effector domain mutants in which the two region s of flexibility were randomized. Secondly, the functional roles of th e minimal Rex effector domain and hydrophobic substitutions were indep endently confirmed by demonstrating that these effector domains could serve as nuclear export signals when conjugated with bovine serum albu min. Nuclear export of the wild-type Rex conjugates was temperature de pendent and sensitive to wheat germ agglutinin and was blocked by a 20 -fold excess of unlabeled conjugates. Together, these studies reveal t hat position-variable hydrophobic interactions within the HTLV-1 Rex e ffector domain mediate nuclear export function.