Embryonic stem (ES) cells serve as a potentially inexhaustible source for tissue replacement in regenerative medicine due to their capability of unlimited self-renewal and multi-lineage differentiation. Vital cellular functions of ES cells require the coordinated action of a large number of proteins that assemble into an array of multi-protein complexes of distinct composition and structure (protein-protein interactions). In addition, physical interactions between regulatory pluripotency transcription factors and their target genes (protein-DNA interactions) provide insights into differential gene expression dictating the pluripotency program. Analysis of protein complexes encompassing intricate protein-protein and regulatory protein-DNA interactions is key to understanding stem cell pluripotency.
Recently, we tested the utility of in vivo biotinylation of transcription factors in mouse ES cells, and have established an in vivo biotinylation system for BirA-mediated specific biotinylation of critical pluripotency factors in mouse ES cells. We developed and optimized an approach for affinity purification of pluripotency protein complexes involving streptavidin capture of biotinylated proteins (dubbed bioSAIP) and demonstrated the feasibility of in vivo biotinylation for mapping global/chromosomal targets of many different transcription factors (dubbed bioChIP-chip) (see Figure 1). Utilizing the technologies we developed, we have constructed a protein interaction network surrounding the pluripotency factor Nanog in mouse ES cells (Wang et al., Nature 2006) and mapped an extended transcriptional network for pluripotency of mouse ES cells (Kim et al. Cell 2008). The network is highly enriched for factors known to be critical in ES cell biology and appears to function as a module for pluripotency. Pluripotency is maintained by many transcription factors that form a highly interconnected protein interaction network including the two homeobox proteins Nanog and Oct4, and a battery of associated proteins of known and unknown functions linking to multiple co-repressor pathways (Figure 2).
Further dissection of the pluripotency network in human ES cells (and induced pluripotent stem cells) and understanding molecular function of the novel factors should illuminate fundamental properties of stem cells and the process of cellular reprogramming, and ultimately lead to precise manipulation and realization of the full clinical therapeutic benefits of these unique cells. Therefore, my lab will be focusing on the following three research areas:
1. Defining protein-protein and protein-DNA interaction networks for pluripotency of human ES cells (and human iPS cells);
2. Dissecting molecular action of novel pluripotency factors on stem cell self-renewal and pluripotency;
3. Elucidating functional significance of novel pluripotency factors in early development and somatic cell reprogramming.
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Faiola, F., Saunders, A., Dang, B., and Wang, J. (2013). An improved in vivo biotinylation strategy combined with FLAG and antibody based approaches for affinity purification of protein complexes in mouse embryonic stem cells. Methods in Molecular Biology, accepted.
Saunders, A., Faiola, F., and Wang, J. (2013). Pursuing self-renewal and pluripotency with the stem cell factor Nanog. Stem Cells 2013 Jul;31(7):1227-36. PMID:23653415.
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McArthur B.D., Sevilla, A., Lenz, M., Muller, F.-J., Schuldt, B.M., Schuppert, A.A., Ridden, S.J., Stumpf, P.S., Fidalgo, M., Ma'ayan, A., Wang, J., and Lemischka, I.R. (2012). Nanog-dependent feedback loops regulate murine embryonic stem cell heterogeneity. Nat. Cell Biol. 2012 Nov;14(11):1139-47. doi: 10.1038/ncb2603. PMID: 23103910
Fidalgo, M., Faiola, F., Pereira, C.-F., Ding, J., Saunders, A., Gingold, J., Schaniel, C., Lemischka, I.R., Silva, J., and Wang, J. (2012). Zfp281 mediates Nanog autorepression through recruitment of the NuRD complex and inhibits somatic cell reprogramming. Proc. Natl. Acad. Sci. USA 109(40), 16202-16207. PMID: 22988117.
Ando K., Kernan, J.L., Liu, P.H., Sandra, T., Logette, E., Tschopp, J., Look, A. T., Wang, J., Bouchier-Hayes, L., and Sidi, S. (2012). PIDD death-domain phosphorylation by ATM determines prodeath versus prosurvival PIDDosome. Mol Cell 47, 681-693. PMID: 22854598.
Yap, K. L., Fraley, S.I., Thiaville, M.M., Jinawath, N., Nakayama, K., Wang, J., Wang, T.L., Wirtz, D., Shih, I.M. (2012). NAC1 is an actin-binding protein that is essential for effective cytokinesis in cancer cells. Cancer Res. 72, 4085–4096. PMID: 22761335.
Wang J. (2012). Deciphering protein complexes and protein interaction networks for stem cell pluripotency. New Frontiers of Network Analysis in Systems Biology (Edited by A. Ma'ayan and B. MacArthur), Chapter 6, 97-118.
Ding, J., Xu, H., Faiola, F., Ma’ayan, A., and Wang, J. (2012). Oct4 links multiple epigenetic pathways to the pluripotency network. Cell Research 22: 155-167. PMID: 22083510.