Transport of BPV-1 virus from the cell membrane to the nucleus was studied in vitro in CV-1 cells. At reduced temperature (4°C), BPV-1 binding to CV-1 cells was unaffected but there was no transport of virions across the cytosol. Electron microscopy showed BPV-1 virions in association with microtubules in the cytoplasm, a finding confirmed by co-immunoprecipitation of L1 protein and tubulin. Internalization of virus was unimpaired in cells treated with the microtubule-depolymerizing drug nocodazole but virions were retained in cytoplasmic vesicles and not transported to the nucleus. We conclude that a microtubule transport mechanism in CV-1 cells moves intact BPV-1 virions from the cell surface to the nuclear membrane.