1. ¹Department of Biochemistry, McGill University, McIntyre Medical Building, 3655 Promenade Sir William Osler, Montreal, Quebec, Canada H3G 1Y6
2. ²Centre de recherche en cancérologie de l'Université Laval, L'Hôtel-Dieu de Québec, CHUQ, Québec, Canada G1R 2J6
3. ³Department of Oncology, McGill University, McIntyre Medical Building, 3655 Promenade Sir William Osler, Montreal, Quebec, Canada H3G 1Y6
4. ⁴McGill Cancer Centre, McGill University, McIntyre Medical Building, 3655 Promenade Sir William Osler, Montreal, Quebec, Canada H3G 1Y6

Correspondence: PE Branton, E-mail: philip.branton@mcgill.ca

Received 17 December 2004; Revised 25 May 2004; Accepted 9 June 2004; Published online 30 August 2004.

Abstract

The adenovirus E4orf4 protein induces p53-independent death of human cancer cells by a mechanism requiring interactions with the B subunit of protein phosphatase 2A. When expressed alone E4orf4 localizes predominantly in the nucleus, although significant levels are also present in the cytoplasm. While tyrosine phosphorylation of E4orf4 and recruitment of Src have been linked with E4orf4 cytoplasmic cell death functions, little is known about the functions of E4orf4 in the nucleus. In this study, we identified an arginine-rich motif (E4ARM; residues 66–75) that is necessary and sufficient for nuclear and nucleolar localization. This motif, which is highly homologous to the arginine-rich nuclear and nucleolar localization motif of some lentiviral proteins, was shown to target heterologous proteins to the nucleus and to nucleoli, functions found to be dependent on the overall charge of the motif rather than on specific residues. Furthermore, mutation of arginine residues to alanines but not to lysines in E4ARM was shown to block such targeting activity and, when introduced into full-length E4orf4, to decrease induction of cell death. Finally, coexpression of the ARM motifs of E4orf4, HIV-1 Tat or Rev along with full-length E4orf4 was seen to decrease E4orf4-dependent cell killing. Thus it appears that targeting of E4orf4 to the nucleus and cell nucleoli by E4ARM is an important component of E4orf4-induced cell death.