HCF-dependent nuclear import of VP16.
Transactivation by VP16 requires the formation of a multicomponent complex, the TAATGAAAT recognition factor complex (TRF.C), that contains in addition to VP16, two cellular proteins, Oct-1 and HCF. HCF binds directly to VP16 and this promotes subsequent interaction of the VP16-HCF complex with the POU DNA-binding domain of Oct-1 and selective assembly onto target sites. Here we demonstrate a novel role of HCF in the intracellular compartmentalization of VP16. We show that while VP16 does not contain a consensus nuclear localization signal (NLS) and is largely cytoplasmic, co-expression with HCF resulted in VP16 nuclear accumulation. A candidate NLS within the C-terminus of HCF was identified and insertion of this motif into green fluorescent protein (GFP) promoted nuclear accumulation. Conversely, removal of this signal from HCF (HCFDeltaNLS) resulted in its cytoplasmic accumulation. Co-expression of HCFDeltaNLS with wild-type (wt) VP16, or of wt HCF with VP16 mutants lacking HCF-binding activity failed to promote the nuclear enrichment of VP16. These results indicate that in addition to its role in stabilizing TRF.C, HCF acts as a nuclear import factor for VP16.
La Boissière
S
S
Marie Curie Research Institute, The Chart, Oxted, Surrey RH8 OTL, UK.
Hughes
T
T
O'Hare
P
P
D000595:Amino Acid Sequence; D000818:Animals; D001693:Biological Transport, Active; D019556:COS Cells; D002451:Cell Compartmentation; D002467:Cell Nucleus / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D017405:Herpes Simplex Virus Protein Vmw65 / Q000378:metabolism*; D051863:Host Cell Factor C1; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D019913:Nuclear Localization Signals; D050811:Octamer Transcription Factor-1; D013347:Subcellular Fractions / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014709:Vero Cells
D004268:DNA-Binding Proteins; C497692:HCFC1 protein, human; D017405:Herpes Simplex Virus Protein Vmw65; D051863:Host Cell Factor C1; D019913:Nuclear Localization Signals; D050811:Octamer Transcription Factor-1; C494165:POU2F1 protein, human; D014157:Transcription Factors