The long terminal repeat-containing retrotransposon Tf1 possesses amino acids in gag that regulate nuclear localization and particle formation.
Tf1 is a long terminal repeat-containing retrotransposon of Schizosaccharomyces pombe that is studied to further our understanding of retrovirus propagation. One important application is to examine Tf1 as a model for how human immunodeficiency virus type 1 proteins enter the nucleus. The accumulation of Tf1 Gag in the nucleus requires an N-terminal nuclear localization signal (NLS) and the nuclear pore factor Nup124p. Here, we report that NLS activity is regulated by adjacent residues. Five mutant transposons were made, each with sequential tracts of four amino acids in Gag replaced by alanines. All five versions of Tf1 transposed with frequencies that were significantly lower than that of the wild type. Although all five made normal amounts of Gag, two of the mutations did not make cDNA, indicating that Gag contributed to reverse transcription. The localization of the Gag in the nucleus was significantly reduced by mutations A1, A2, and A3. These results identified residues in Gag that contribute to the function of the NLS. The Gags of A4 and A5 localized within the nucleus but exhibited severe defects in the formation of virus-like particles. Of particular interest was that the mutations in Gag-A4 and Gag-A5 caused their nuclear localization to become independent of Nup124p. These results suggested that Nup124p was only required for import of Tf1 Gag because of its extensive multimerization.
Kim
Min-Kyung
MK
Section on Eukaryotic Transposable Elements, Laboratory of Gene Regulation and Development, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.
Claiborn
Kathryn C
KC
Levin
Henry L
HL
D021581:Active Transport, Cell Nucleus; D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000502:physiology*; D002467:Cell Nucleus / Q000378:metabolism*; D015683:Gene Products, gag / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D019913:Nuclear Localization Signals / Q000502:physiology*; D028861:Nuclear Pore Complex Proteins / Q000235:genetics / Q000378:metabolism; D018626:Retroelements / Q000235:genetics / Q000502:physiology*; D012568:Schizosaccharomyces / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000378:metabolism; D020079:Terminal Repeat Sequences / Q000235:genetics; D014780:Viruses / Q000378:metabolism
D000596:Amino Acids; D015683:Gene Products, gag; D019913:Nuclear Localization Signals; D028861:Nuclear Pore Complex Proteins; D018626:Retroelements; D029702:Schizosaccharomyces pombe Proteins; C120536:nup124 protein, S pombe; D000409:Alanine