Chang-Deng Hu, M.D., Ph.D. Associate Professor of Medicinal Chemistry and Moleculary Pharmacology Phone: (765) 496-1971 Fax: (765) 494-1414 E-mail: hu1@purdue.edu Personal webpage: http://people.pharmacy.purdue.edu/~hu1/ Full directory listing

Specialization: AP-1 in Cancer and C. elegans Development

Education

Research: AP-1 in Cancer and C. elegans Development

Protein-protein interactions are essential for transmitting extracellular signals into cells and for coordinating cellular functions. Although many interaction maps have been generated over the past few years using genome-wide approaches, such as yeast two-hybrid and proteomics, it remains a challenge to prove these interactions in vivo. We have developed a novel bimolecular fluorescence complementation (BiFC) assay to directly visualize protein-protein interactions in living cells (Molecular Cell, 9, 789-798, 2002). This assay is based on the formation of a bimolecular fluorescent complex between two halves of YFP (yellow fluorescent protein) fused to a pair of interaction partners. To study how each protein selects its interacting partners in response to specific signals, we have taken advantage of spectral variants of green fluorescent protein and further established a multicolor bimolecular fluorescence complementation (multicolor BiFC) assay (Nature Biotechnology, 21, 539-545, 2003). The multicolor BiFC assay allows us to study multiple protein interactions simultaneously in the same cell. Recently, the identification of several fluorescent protein fragments derived from the new fluorescent proteins, Venus, Citrine and Cerulean, has further expanded our capability to analyze protein-protein interactions under physiological conditions (BioTechniques, 40, 61-66, 2006).

AP-1 in cancer: Activator protein 1 (AP-1) belongs to the basic region leucine zipper (bZIP) family of transcription factors and functions as homodimers or heterodimers formed among the members of Fos, Jun, ATF2 and Maf family of proteins to regulate gene expression. AP-1 activity can be induced by both physiological stimuli and environmental stresses, thereby regulating a wide range of cellular processes including cell proliferation, differentiation, death, and stress responses. Deregulated AP-1 activity is implicated in many human diseases including cancer. Furthermore, AP-1 proteins also interact with many other transcriptional regulatory proteins, such as the Rel family, SMADs family, hormone receptors, and coactivators CBP/p300. These cross-family interactions further increase the complexity of the regulation of target genes. To study how the interactions of AP-1 proteins with those within, as well as across the families determine cellular responses, we are using our BiFC assays, in combination with molecular, cellular, biochemical, genetic and systems biology approaches, to visualize these interactions in living cells and to investigate the regulation and functional consequences of the interactions. Our current projects include:

1. Regulation and function of ATF2 subcellular localization.
2. Regulation and implication of neuroendocrine differentiation in prostate cancer progression and therapy.
3. Role of cross-family interactions of NF-κB with AP-1 in chemoresistance and radioresistance.
4. Screening of small molecules for the inhibition of protein-protein interactions using a multicolor BiFC-based HTS system.

AP-1 in C. elegans development: Gene targeting has been widely used to study the functions of genes in vivo. However, problems often encountered using gene knock-out studies are embryonic lethality or lack of obvious phenotypes. The former prevents further evaluation of the targeted genes during the entire developmental process and the latter frequently reflects functional redundancy of homologous genes or isoforms. Because AP-1 functions as heterodimers or homodimers, the regulation of dimer formation plays a pivotal role in the control of their transcriptional activities. Accordingly, monitoring their interactions throughout development will provide a substantial link to the roles of AP-1 in development. The establishment of the BiFC assays has endowed us with a unique way to study protein-protein interactions in living animals. We are applying the BiFC assays to study the temporal and spatial interactions of C. elegans AP-1 proteins in living worms.

To learn more about our research, please visit our lab homepage at http://people.pharmacy.purdue.edu/~cdhu/.

Representative Publications

Hiatt, S.M., Duren, H.M. Shyu, Y., Ellis, R.E., Hisamoto, N., Matsumoto, K., Kariya, K., Kerppola, T.K., and Hu, C.D.. C. elegans FOS-1 and JUN-1 regulate plc-1 expression to control ovulation. Mol. Biol. Cell 20:3888-3895 (2009)

Yuan, Z., Gong, S., Song, B., Mei, Y., Hu, C., Li, D., Thiel, G., Hu, C.D., and Li, M. Opposing role for ATF2 and c-Fos in c-Jun-mediated apoptosis induced by potassium deprivation in cerebellar granule neurons. Mol. Cell. Biol. 29:2431-2442 (2009)

Deng, X., Liu, H., Huang, J., Cheng, L., Keller, E.T., Parsons, S.J., and Hu, C.D. Ionizing radiation induces prostate cancer neuroendocrine differentiation through interplay of CREB and ATF2: Implications for disease progression. Cancer Res. 68:9663-9670 (2008)

Shyu, Y. and Hu, C.D. Fluorescence complementation: an emerging tool for biological research. Trends Biotechnol. 26:622-630 (2008)

Shyu, Y., Suarez, C., and Hu, C.D. Visualization of ternary complexes in living cells by using a BiFC-based FRET analysis. Nat. Protoc. 3:1693-1702 (2008)

Vidi, P.A., Chemel, B.R., Hu, C.D., Watts, V.J. Ligand-Dependant Oligomerization of Dopamine D₂ and Adenosine A_2A Receptors in Living Neuronal Cells. Mol. Pharmacol. 74:544-551 (2008)

Hiatt, S.M., Shyu, Y., Duren, H.M, and Hu, C.D. Bimolecular fluorescence complementation (BiFC) analysis of protein interactions in living C. elegans. Methods, 45:185-191 (2008)

Shyu, Y., Fox, SM., Duren, HM., Ellis, R.E., Kerppola, T.K. and Hu, C.D. Visualization of protein interaction in living Caenorhabditis elegans using bimolecular fluorescence complementation (BiFC) analysis. Nat. Protoc., 4:588-596 (2008)

Shyu, Y., Suarez, C., and Hu, C.D. Visualization of AP-1-NF-κB ternary complexes in living cells by using a BiFC-based FRET. Proc. Natl. Acad. Sci. U.S.A., 105:151-156 (2008)

Liu, H., Deng, X., Shyu, Y., Li, J.J., Taparowsky, EJ., and Hu, C.D. Mutual regulation of c-Jun and ATF2 by transcriptional activation and subcellular localization. The EMBO J., 25:1058-1069 (2006).