[An efficient high-throughput screening assay against nuclear transport].
Nuclear transport, an essential cellular process of eukaryotic cells, plays critical roles in differentiation , development, as well as viral disease and oncogenesis, making the small molecules that target this process very intriguing not only in fundamental research but also in disease treatment. However, only one compound, Leptomycin B, is commercially available to inhibit nuclear transport. Therefore, it will be a great advantage to establish an assay that targets the whole nuclear transport pathway for screening and obtaining small molecule that regulates nuclear transport. In this study, we established an efficient nuclear transport assay based on the reconstitution of GFP by NZGFP and CZGFP. We constructed NZGFP-NES and CZGFP-NLS, making them locate in cytoplasm and nucleus separately. Their distribution will be changed when nuclear transport is interfered, resulting in co-localization of NZGFP-NES and CZGFP-NLS and subsequent reconstitution of fluorescent GFP. The inhibiting effect of Leptomycin B on nuclear transport can be sensitively detected by NZGFP-NES/CZGFP-NLS report system, providing an efficient assay for high-throughput screening of small molecule against nuclear transport.
ShenzhenCenter for Disease Control and Prevention, Shenzhen 518055, China. luomin4653@163.net
Min
M
Luo
Quan-Cang
QC
Zhang
Zhi-Gang
ZG
Lu
D021581:Active Transport, Cell Nucleus / Q000187:drug effects*; D000595:Amino Acid Sequence; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D023061:Gene Order; D017930:Genes, Reporter; D006367:HeLa Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D008969:Molecular Sequence Data
D005231:Fatty Acids, Unsaturated; C038753:leptomycin B