Definition of a consensus transportin-specific nucleocytoplasmic transportsignal

The low cytoplasmic and high nuclear concentration of the GTP-bound form of Ran provides directionality for both nuclear protein import and export. Bo th import and export factors bind RanGTP directly, yet this interaction pro duces opposite effects; in the former case, RanGTP binding induces nuclear cargo release, whereas in the latter, RanGTP binding induces nuclear cargo assembly. Therefore, nuclear import and export receptors and their protein recognition sites are predicted to be distinct. Nevertheless, the similar t o 38-amino acid M9 sequence present in heterogeneous nuclear ribonucleoprot ein Al has been reported to serve as both a nuclear localization signal and a nuclear export signal, even though only one protein, the nuclear import factor transportin, has been shown to bind M9 directly. We have used a comb ination of mutational randomization followed by selection for transportin b inding to exhaustively define amino acids in M9 that are critical for trans portin binding in vivo, As expected, the resultant similar to 12-amino acid transportin-binding consensus sequence is also predictive of nuclear local ization signal activity. Surprisingly, however, this extensive mutational a nalysis failed to dissect M9 nuclear localization signal and nuclear export signal function. Nevertheless, transportin appears unlikely to be the M9 e xport receptor, as RanGTP can be shown to block M9 binding by transportin n ot only in vitro, but also in the nucleus in vivo, This analysis therefore predicts the existence of a nuclear export receptor distinct from transport in that nevertheless shares a common protein-binding site on heterogeneous nuclear ribonucleoprotein A1.