Sequence requirements for the nuclear localization of the murine cytomegalovirus M44 gene product pp50

Virology. 1999 Jun 20;259(1):43-59. doi: 10.1006/viro.1999.9700.

Abstract

The murine cytomegalovirus (MCMV) M44 gene product pp50 is normally present in the nuclei of virus-infected cells. During transient expression of pp50 in COS-1 cells, the phosphoprotein was readily detectable in the nuclei, indicating that it possesses a nuclear localization signal (NLS). Studies on the subcellular locations of N- and C-terminal deletion mutants of pp50 suggested that alterations in both the C terminus and the highly conserved N-terminal domains of pp50 affect nuclear localization. In particular, the C-terminal 11 amino acids of pp50, which includes a "KKQK" motif, were able to mediate the import of a beta-galactosidase fusion protein into the nucleus. The pair of lysine residues in this motif constitutes an essential element of the C-terminal NLS as mutation of this motif to AAQK directly affected the nuclear localization of either pp50 or beta-galactosidase fusion proteins containing the C-terminal portion of pp50. Furthermore our results indicated that the functionality of the C-terminal NLS is dependent on the structural integrity of the highly conserved N-terminal portion of the molecule, as deletion of amino acids 157-201 alone adversely affected nuclear localization. In the absence of a functional C-terminal NLS, the subcellular localization of pp50 is sensitive to potential conformational changes induced by mutations within the N-terminal half of the molecule. Under those circumstances, mutation of the YK residues at position 22-23 or deletion of amino acids 267-283 was sufficient to produce a protein that was impaired in nuclear import or retention.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites / genetics
  • Cell Nucleus / virology
  • DNA-Binding Proteins / genetics*
  • Genes, Viral
  • Mice
  • Molecular Sequence Data
  • Muromegalovirus / genetics*
  • Sequence Analysis
  • Sequence Deletion
  • Viral Nonstructural Proteins / genetics*

Substances

  • DNA-Binding Proteins
  • Viral Nonstructural Proteins
  • phosphoprotein 50, cytomegalovirus