Human cytomegalovirus pp65 lower matrix phosphoprotein harbours two transplantable nuclear localization signals

1 Istituto di Genetica Biochimica ed Evoluzionistica, Consiglio Nazionale delle Ricerche, via Abbiategrasso 207, I-27100 Pavia and 2 Virology Unit, Istituto di Clinica delle Malattie Infettive, IRCCS Policlinico San Matteo, I-27100 Pavia, Italy Human cytomegalovirus phosphoprotein pp65 is targeted to the cell nucleus immediately after infection. Deletion and point mutation analysis of the pp65 gene expressed in insect cells showed that two hydrophilic regions (HP1 and HP2) within the pp65 C-terminal 40% each harboured an independent nuclear localization signal (NLS); strong association to the nuclear stroma also requires the N-terminal domain. Either region, when fused to chloramphenicol acetyltransferase, localized the reporter protein to the nucleus in insect cells as well as in NIH 3T3 cells and human lung fibroblasts. In addition, HP1 was found to be the target of pp65 Ser/Thr phosphorylation in insect cells and a prokaryotically expressed HP1 was actively phosphorylated in vitro by casein kinase II, for which two site clusters map in HP1. These findings indicate that pp65 includes two NLSs, one of which has the potential to be modulated by phosphorylation. * Author for correspondence. Fax +39 382 422286. e-mail gallina@ipvgbe.igbe.pv.cnr.it Received 29 September 1995; accepted 12 January 1996.