Premium
Influence of Serine O ‐Glycosylation or O ‐Phoshorylation Close to the vJun Nuclear Localisation Sequence on Nuclear Import
Author(s) -
Schlummer Stefanie,
Vetter Robin,
Kuder Norman,
Henkel Andreas,
Chen YongXiang,
Li YanMei,
Kuhlmann Jürgen,
Waldmann Herbert
Publication year - 2006
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.200500212
Subject(s) - nls , nuclear localization sequence , nuclear transport , importin , serine , phosphorylation , chemistry , glycosylation , biochemistry , cell nucleus , cytoplasm , gene
Nuclear import triggered by the nuclear‐localisation sequence (NLS) of the viral Jun (vJun) protein is mediated by phosphorylation of a serine close to the NLS. Since phosphorylation and glycosylation of serine residues are often in a reciprocal “yin–yang” relationship, we investigated whether glycosylation of this serine with O‐linked N ‐acetylglucosamine ( O ‐GlcNAc) would also regulate nuclear import via the vJun NLS. Peptides containing the vJun NLS with an adjacent O ‐phosphorylated, O ‐GlcNAc‐functionalised or unmodified serine, and equipped with an N‐terminal biotin or a 7‐nitrobenz‐2‐oxa‐1,3‐diazolyl (NBD) fluorescent label, were synthesised on the solid phase by means of an Fmoc/Boc strategy and a Pd 0 ‐sensitive HYCRON linker. Fluorescence‐polarisation measurements on the NBD‐labelled peptides indicated that modification with phosphate or O ‐GlcNAc leads to a decrease in affinity to the import‐mediating adapter protein, importin α, of about one order of magnitude compared to the unmodified NLS. Microinjection of biotinylated NLS peptide conjugated with fluorescently labelled avidin into NIH/3T3 and MDCK cells, revealed that avidin–unmodified‐NLS peptide was rapidly imported into the nucleus. However, either phosphate or O ‐GlcNAc next to the NLS caused almost complete exclusion of the protein conjugate from nuclear import. These findings indicate that nuclear import by the vJun NLS might not be regulated by a “yin–yang” modification of an adjacent serine with phosphate or O ‐GlcNAc. Rather, negative regulation of binding between the polybasic NLS and importin by a negatively charged or a bulky, uncharged residue appears likely.