A computer-assisted two-dimensional gel electrophoresis approach for studying the variations in protein expression related to an induced functional repression of NFκB in lymphoblastoid cell lines

Abstract

Strategies are needed for conclusive interpretation of two-dimensional gel electrophoresis (2-D PAGE) maps in order to identify pertinent differences in protein expression during regulation of the transcription of discrete sets of genes. The model used in this study was a human lymphoblastoid cell line in which a functional repression of the transcription factors NFκB was obtained by induction of overexpression of IκBα, a physiological inhibitor of NFκB. The analytical methodology used relies on the comparison of two sets of 2-D PAGE maps for detecting differences in protein expression between samples overexpressing or not overexpressing IκBα. The analysis was based on a combination of an automatic computerized analysis, constituting an actual aid for deciding, and of an interactive visual validation, corresponding to the interpretation of computer propositions. This strategy is proposed as a rapid way to detect potential variations in protein expression applicable to any biological model. In this study, correspondence analysis data made it possible to discrimate between the samples overexpressing or not overexpressing IκBα, and pointed out some of the potential meaningful spots characterizing the samples in which NFκB was active. Then, after visual validation of the computer data, 53 polypeptides were considered to be different in the two classes of gels. Five polypeptides were specifically found in both samples overexpressing IκBα. The overexpression of IκB also induced a lower expression of 11 polypeptides. Finally, 15 polypeptides were only expressed in samples in which IκBα was not overexpressed and, consequently, in which NFκB factors were active. Thus, these polypeptides are candidates for further analysis as putative target gene products of NFκB.