Marit Otterlei et Per Bruheim: The AlkB homolog 2 PCNA interacting motif, APIM, interacts with PCNA and the beta-clamp; roles of PCNA in regulation of cellular signalling and potential applications of APIM-peptides as anti-cancer and antibacterial agents
Organisateur : Vincent Géli
Lieu : IPC2
Norwegian University of Science and Technology, Trondheim, Norway
The two PCNA-interacting motifs, AlkB homolog 2 PCNA interacting motif, APIM, and PCNA-interacting protein-box, PIP-box, are present in ~ 500 proteins (1). Many of the APIM-containing proteins are proteins involved in cellular stress responses, including multiple proteins in cell signaling. Targeting PCNA with a cell-penetrating peptide containing APIM (APIM-peptide) was previously shown to affect PI3K/AKT-signaling, and to induce apoptosis in a dose-dependent and cell cycle independent manner (2,3). After extensive metabolome (4,5), kinome/proteome (6), and transcriptome profiling assays, we have recently found that targeting PCNA affects multiple signaling pathways in multiple cell lines. This leads to reduced primary metabolism in MM cell lines while less or no effects were observed in other cell lines examined. Our data suggest cell-specific roles for PCNA as a scaffold protein in cellular signaling.
Interestingly, APIM-peptides also have antibacterial and anti-mutagenic activities. PCNA and the β-clamp belong to the structurally conserved DNA sliding clamp family, and both contain a hydrophobic pocket in their C-terminal domain mediating multiple protein-protein interactions via specific binding motifs. Both APIM and PIP-box, bind to this site on PCNA, and a broadly defined 5-6 amino acids clamp binding motif (CBM), found on prokaryotic DNA polymerases and other β-clamp interacting proteins, mediates interaction with this site on the β-clamp. Our data suggest that APIM-peptides bind to the β-clamp and inhibits interactions multiple proteins.
1. Gilljam, K.M., Feyzi, E., Aas, P.A., Sousa, M.M., Müller, R., Vågbø, C.B., Catterall, T.C., Liabakk, N.B., Slupphaug, G. and Drabløs, F. (2009) Identification of a novel, widespread, and functionally important PCNA-binding motif. The Journal of cell biology, 186, 645-654.
2. Olaisen, C., Müller, R., Nedal, A. and Otterlei, M. (2015) PCNA-interacting peptides reduce Akt phosphorylation and TLR-mediated cytokine secretion suggesting a role of PCNA in cellular signaling. Cellular signalling, 27, 1478-1487.
3. Müller, R., Misund, K., Holien, T., Bachke, S., Gilljam, K.M., Våtsveen, T.K., Rø, T.B., Bellacchio, E., Sundan, A. and Otterlei, M. (2013) Targeting proliferating cell nuclear antigen and its protein interactions induces apoptosis in multiple myeloma cells. PloS one, 8, e70430.
4. Kvitvang, H.F., Kristiansen, K.A. and Bruheim, P. (2014) Assessment of capillary anion exchange ion chromatography tandem mass spectrometry for the quantitative profiling of the phosphometabolome and organic acids in biological extracts. Journal of chromatography. A, 1370, 70-79.
5. Lien, S.K., Sletta, H., Ellingsen, T.E., Valla, S., Correa, E., Goodacre, R., Vernstad, K., Borgos, S.E.F. and Bruheim, P. (2013) Investigating alginate production and carbon utilization in Pseudomonas fluorescens SBW25 using mass spectrometry-based metabolic profiling. Metabolomics, 9, 403-417.
6. Petrovic, V., Olaisen, C., Sharma, A., Nepal, A., Bugge, S., Sundby, E., Hoff, B.H., Slupphaug, G. and Otterlei, M. (2017) On-column trypsinization allows for re-use of matrix in modified Multiplexed Inhibitor Beads assay. Analytical Biochemistry, 523, 10-16.