Background: A hallmark of retroviral gene expression is that unspliced retroviral genomic RNA is exported to the cytoplasm, whereas endogenous intron-containing cellular RNAs are usually retained in the nucleus. In complex retroviruses, such as human immunodeficiency virus-1 (HIV-1), nuclear export is accomplished by the interaction of a virally encoded protein, Rev, with a cis-acting RNA element, the Rev-responsive element (RRE). In type D retroviruses, such as the simian retrovirus type 1 (SRV-1), however, genomic RNA is exported by cellular factor(s) that interact with a conserved cis-acting RNA element, the constitutive transport element (CTE). Results: We found that the CTE was exported in a specific and saturable fashion from Xenopus oocyte nuclei. When inserted into the intron of an adenovirus-derived pre-mRNA, the CTE did not affect splicing efficiency but promoted the nuclear export of the excised intron lariat that is normally retained within the nucleus. Export of CTE-containing RNAs to the cytoplasm was not affected by the heterogeneous nuclear ribonucleoprotein A1 or an excess of peptides corresponding to the Rev nuclear export signal. Microinjection of saturating amounts of CTE RNA did not affect tRNA export or Rev-mediated export but did inhibit mRNA export. CTE-mediated export was found to be dependent on Ran-mediated GTP hydrolysis. Conclusion: The Rev–RRE system and the CTE direct intron-containing RNAs to distinct export pathways. Although previous data have suggested that Rev uses the same export pathway as uracil-rich small nuclear RNAs and 5S ribosomal RNA, the CTE seems to interact with evolutionarily conserved factors that are essential for cellular mRNA export.