Human cytomegalovirus pp65 lower matrix phosphoprotein harbours two transplantable nuclear localization signals.
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.
Gallina
A
A
Instituto di Genetica Biochimica ed Evoluzionistica, Consiglio Nazionale delle Ricerche, Pavia, Italy. gallina@ipvgbe.igbe.pv.cnr.it
Percivalle
E
E
Simoncini
L
L
Revello
M G
MG
Gerna
G
G
Milanesi
G
G
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D003001:Cloning, Molecular; D003587:Cytomegalovirus / Q000378:metabolism*; D006801:Humans; D008168:Lung; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016366:Open Reading Frames; D010750:Phosphoproteins / Q000096:biosynthesis / Q000378:metabolism*; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D017384:Sequence Deletion; D016324:Sequence Tagged Sites; D015398:Signal Transduction; D018411:Spodoptera; D014162:Transfection; D014763:Viral Matrix Proteins / Q000096:biosynthesis / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D010750:Phosphoproteins; D011994:Recombinant Proteins; D014763:Viral Matrix Proteins; C065418:cytomegalovirus matrix protein 65kDa; D015500:Chloramphenicol O-Acetyltransferase