Characterization of the nuclear targeting signal of REST/NRSF.
RE-1 silencer transcription factor (REST), also known as neuron-restrictive silencer factor (NRSF), contains nine Cys2-His2 type zinc finger domains (ZFDs). REST/NRSF is localized to the nucleus, where it represses the transcriptional activity of a large number of neuronal genes in non-neuronal cells. It has been suggested that REST/NRSF contains a nuclear localization signal (NLS) corresponding to amino acids (512-522). However, our studies showed that REST4, a REST/NRSF splicing isoform, which contains the N-terminal 5 of 9 ZFDs, efficiently localized to the nucleus. On the other hand REST1, another REST/NRSF splicing isoform, which contains 4 of the 9 ZFDs, localized to the cytosol. In this study REST-DeltaC, which contains 8 ZFDs with the NLS (512-522) deleted, was found to localize to the nucleus in HeLa, COS and PC12 cells. Complete deletion or mutation of NLS (512-522) still permitted REST/NRSF to be localized to the nucleus in HeLa, COS and PC12 cells. In contrast REST/NRSF constructs which contain a deletion of ZFD-5 mislocalized to the cytosol. A point mutation in the zinc finger structure that disrupts its conformation remains nuclear. These data suggest that REST/NRSF contains a NLS around ZFD-5, while the putative NLS at residues 512-522 is non-functional.
Shimojo
Masahito
M
Department of Molecular and Cellular Biochemistry, University of Kentucky College of Medicine, Biomedical Biological Sciences Research Building, 741 Limestone Street, Lexington, KY 40536-0509, USA. mshim1@uky.edu
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus; D002522:Chlorocebus aethiops; D003600:Cytosol / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D016716:PC12 Cells; D017354:Point Mutation; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D021382:Protein Sorting Signals; D021381:Protein Transport; D051381:Rats; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D017384:Sequence Deletion; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D016335:Zinc Fingers; D000099229:RE1-Silencing Transcription Factor
D020033:Protein Isoforms; D021382:Protein Sorting Signals; D012097:Repressor Proteins; D014157:Transcription Factors; D000099229:RE1-Silencing Transcription Factor