Nuclear localization of oligonucleotides decoy effect on nuclear factor-kappaB activity.
- Author:
Yingxun LIU
1
;
Fusheng QUAN
;
Jinke WANG
;
Xueyao BAI
Author Information
1. College of Veterinary Medicine, Northwest A&F University, Yangling 712100, China.
- Publication Type:Journal Article
- MeSH:
Base Sequence;
Cell Nucleus;
metabolism;
HeLa Cells;
Humans;
Molecular Sequence Data;
NF-kappa B;
genetics;
metabolism;
Nuclear Localization Signals;
genetics;
Oligonucleotides;
genetics;
metabolism;
Transfection
- From:
Chinese Journal of Biotechnology
2010;26(12):1683-1689
- CountryChina
- Language:Chinese
-
Abstract:
To investigate the effect of the localization of oligonucleotides decoy (ODNs decoy) on the activation of nuclear factor-kappaB (NF-kappaB) in TNF-alpha induced HeLa cells. The mercapto group-modified nuclear localization signal (NLS) peptide was covalently conjugated to amino group-modified NF-kappaB ODNs decoy by Sulfo-SMCC cross-linker. The NLS-ODNs decoy was transfected into HeLa cells by TransME transfection reagent. The intracellular distribution of fluorescent labeled NLS-ODNs decoy was detected with a microscope. The cell viability was detected by MTT assay, and then the activity of NF-kappaB in cell nuclear extract was assayed by electrophoretic mobility shift assay (EMSA). The results showed that NLS peptide was successfully conjugated to ODNs decoy by Sulfo-SMCC cross-linker. The NLS-ODNs decoy effectively entered into nucleus with high rate of 17.9%. It was observed that the cell viability of HeLa cell was not significantly affected by the transfection of NLS-ODNs decoy, while NLS-ODNs decoy significantly inhibited the activation of NF-kappaB in TNF-alpha induced HeLa cells nuclear extracts. This experiment can provide a new covalent conjugation of NLS peptide to ODNs can effectively drive decoy into nucleus, and thus improve its inhibitory effects on the activation a transcription factor.
