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A Protein Kinase CK2 Site Flanking the Nuclear Targeting Signal Enhances Nuclear Transport of Human Cytomegalovirus ppUL44
Author(s) -
Alvisi Gualtiero,
Jans David A.,
Guo Jinjin,
Pinna Lorenzo A.,
Ripalti Alessandro
Publication year - 2005
Publication title -
traffic
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.677
H-Index - 130
eISSN - 1600-0854
pISSN - 1398-9219
DOI - 10.1111/j.1600-0854.2005.00331.x
Subject(s) - nuclear transport , nuclear localization sequence , biology , importin , nuclear protein , nuclear export signal , microbiology and biotechnology , nls , cell nucleus , biochemistry , transcription factor , gene , cytoplasm
The processivity factor of the human cytomegalovirus (HCMV) DNA polymerase phosphoprotein ppUL44 plays an essential role in viral replication, showing nuclear localization in infected cells. The present study examines ppUL44's nuclear import pathway for the first time, ectopic expression of ppUL44 revealing a strong nuclear localization in transfected COS‐7 and other cell types, implying that no other HCMV proteins are required for nuclear transportation and retention. We show that of the two potential nuclear localization signals (NLSs) located at amino acids 162–168 (NLS1) and 425–431 (NLS2), NLS2 is necessary and sufficient to confer nuclear localization. Moreover, using enzyme‐linked immunosorbent assays and gel mobility shift assays, we show that NLS2 is recognized with high affinity by the importin (IMP) α/β heterodimer. Using gel mobility shift and transient transfection assays, we find that flanking sequences containing a cluster of potential phosphorylation sites, including a consensus site for protein kinase CK2 (CK2) at Ser 413 upstream of the NLS, increase NLS2‐dependent IMP binding and nuclear localization, suggesting a role for these sites in enhancing UL44 nuclear transport. Results from site‐directed mutagenic analysis and live‐cell imaging of green fluorescent protein (GFP)‐UL44 fusion protein‐expressing cells treated with the CK2‐specific inhibitor 4,5,6,7‐tetrabromobenzotriazole are consistent with phosphorylation of Ser 413 enhancing ppUL44 nuclear transport.