The 605 amino acid E1 protein of bovine papillomavirus type 1 (BPV-1) is a 
multifunctional nuclear protein required for viral DNA replication. A nuclear 
localization signal (NLS) sequence was previously defined by point mutations in 
three short adjacent clusters of basic amino acids located in the amino-terminal 
region of the E1 protein. In this study, we used a fusion protein approach to 
evaluate the contribution of other regions of the E1 protein to nuclear 
transport. The nearly full-length E1 gene and six non-overlapping subfragments 
were each fused in-frame with the lacZ gene in a eukaryotic expression vector. 
Each clone was electroporated into COS-1 cells, and the intracellular location 
of the E1-beta-galactosidase fusion proteins was determined by 
immunofluorescence. Only the constructs containing the full-length E1 or a 
single subregion (E1-259; amino acids 84 to 166) produced fusion proteins that 
entered the nucleus. Point mutations in the NLS sequences of the E1-259-lacZ 
construct prevented nuclear translocation of the corresponding fusion protein. 
This confirms the previous result that the cluster of basic amino acids is 
critical for nuclear transport. Furthermore, the data obtained in this 
investigation indicated that the region of E1 containing the NLS sequence was 
not only necessary, but was also sufficient for nuclear localization. No other 
region of E1 contained independent nuclear localization activity.