Nuclear entry, oligomerization, and DNA binding of the Drosophila heat shock transcription factor are regulated by a unique nuclear localization sequence.
In normally growing Drosophila cultured cells the Drosophila heat shock transcription factor (dHSF) is localized in the cytosol and translocates into the nucleus after heat shock. In the cytosol of nonshocked cells, the dHSF is present as a monomer that cannot bind DNA. Upon stress, the dHSF enters the nucleus where it is observed to be a trimer. A novel nuclear localization sequence (NLS) in the dHSF was found to be responsible for stress-dependent nuclear entry. Deletion of the NLS prevents nuclear entry, as expected, yet surprisingly also allows constitutive oligomerization and DNA binding in the cytosol. Further analysis of the NLS by mutagenesis suggests that the two functions of nuclear entry and oligomerization are separable in that distinct residues present in the NLS are responsible for each. Mutations in certain basic residues completely block nuclear entry, as expected for a constitutive NLS. In addition, two residues were found in the NLS that, when altered, allowed constitutive nuclear entry of dHSF independent of stress. These residues may interact with a putative cellular component or possibly other domains of the HSF to prevent nuclear entry in normally growing cells. The NLS can also function autonomously to target a beta-galactosidase fusion protein into the nucleus in a heat shock-dependent fashion.
Zandi
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E
Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena 91125, USA.
Tran
T N
TN
Chamberlain
W
W
Parker
C S
CS
D000595:Amino Acid Sequence; D000818:Animals; D001704:Biopolymers; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004331:Drosophila melanogaster; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D021382:Protein Sorting Signals / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D001616:beta-Galactosidase / Q000378:metabolism
D001704:Biopolymers; D004268:DNA-Binding Proteins; D000076249:Heat Shock Transcription Factors; D006360:Heat-Shock Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D001616:beta-Galactosidase