Differential proteomic analysis of nuclear extracts from thyroid cell lines

作者：Salzano AM; Paron I; Pines A; Bachi A; Talamo F; Bivi N; Vascotto C; Damante G; Quadrifoglio F; Scaloni A; Tell G^*

来源：Journal of Chromatography B-Analytical Technologies in the Biomedical and Life Sciences, 2006, 833(1): 41-50.

DOI：10.1016/j.jchromb.2005.12.025

• differential proteomics
• thyroid
• differentiation
• APE1/Ref-1
• transcriptional regulation
• nucleus

Nuclear proteins play a major role in controlling cell functions. Differential proteomic analysis of nuclear proteins by combined 2D gel electrophoresis (2D-E) and mass spectrometry procedures can provide useful information to understand the control of cell proliferation and differentiation. To identify proteins involved in dedifferentiation, we used a differential proteomics approach by comparing nuclear extracts from the differentiated rat thyroid cell line FRTL-5 and the derived undifferentiated Ki-mol cell line, obtained by transformation with the Ki-ras oncogene. Thirteen proteins were identified as differently expressed in the nuclear compartment between the two cell lines. RT-PCR analysis performed on seven differently expressed genes showed that only in two cases the difference may be ascribable to a transcriptional mechanism. Since one of the identified proteins, namely apurinic apyrimidinic endonuclease/redox effector factor-1 (APE 1/Ref-1), is suspected to play a role in thyroid tumorigenesis, we used a glutathione S-transferase (GST)-pulldown assay coupled to a 2D electrophoretic/matrix assisted laser desorption ionization-time of flight (MALDI-TOF)-mass spectrometry (MS) analysis to detect and identify its interacting partners. We show here that P-actin directly interacted with APE 1/Ref-1, as confirmed by co-immunoprecipitation assays and that this interaction was enhanced by oxidative stress on FRTL-5 cells.

• 出版日期2006-3-20