Our lab investigates signal transduction pathways that regulate the growth of normal and neoplastic cells. We are interested in multiple aspects of these pathways, including ligand binding, intracellular phosphorylation, gene transcription and the resulting cellular responses that these pathways induce.
One major area of investigation is the interferon-alpha pathway, which is a model for signaling by many cytokines and polypeptide hormones. There are three key classes of molecules involved in this set of signaling pathways: multi-subunit receptors, JAK family tyrosine kinases, and STAT family transcription factors. The receptors are constitutively associated with the JAK kinases, forming so-called "broken" receptors. Ligand binding activates the kinases, which auto-phosphorylate and phosphorylate the receptors. One or more of the receptor phosphorylation sites can then recruit the STATs via their SH2 domains, and thereby facilitate their activation/phosphorylation.
Over the past few years, our lab has been involved in demonstrating that the JAK kinases are activated by cytokines, characterizing the interaction between the kinases and the receptors, identifying STAT docking sites on the interferon receptor and partially reconstituting the pathway using chimeric receptors. Most recently we have discovered that one of the subunits of the interferon-alpha receptor (IFNaR2) can be proteolytically cleaved in a manner similar to the process that regulates signaling by the Notch proteins. Much of our currect work focuses on characterizing this type of signaling, which is known as 'regulated intramembrane proteolysis' (RIP). In a second project, we are also collaborating with Dave Van Vranken, in the Department of Chemistry, to explore the mechanism of action of madindoline, an inhibitor of cytokine signaling.
A second major area of investigation in our laboratory concerns the molecular mechanisms involved in the pathogenesis of human prostate cancer. This cancer is now the most frequent malignant tumor in men in the U.S. These tumors are initially dependent on the presence of androgens to maintain viability, and therefore, most of these cancers can be initially treated by androgen deprivation therapy. However, this treatment ultimately fails because the tumors become androgen independent. Androgen deprivation therapy slows tumor growth by inducing apoptosis (this also occurs in normal prostate epithelial cells). We are exploring the mechanisms of apoptosis in prostate epithelial cells, with a particular emphasis on the role of the death receptor signaling pathway. One of our working hypotheses is that androgen withdrawal inhibits the transcription of FLIP, a negative regulator of this death-signaling pathway, thereby facilitating apoptosis. We also hypothesize that one or more death receptor ligands are required for androgen withdrawal induced apoptosis. We are employing a variety of model systems, including: castration-induced apoptosis of the rodent prostate glands, TGFß-induced apoptosis of two cell lines (rat NRP-152s and human DU-145s), and androgen deprivation of human LNCaP prostate cancer cells.
Lastly, I am also the Associate Director of the Molecular Pathology Laboratory in the Department of Pathology and Laboratory Medicine. This is a clinical lab devoted to the use of modern molecular biologic techniques in the diagnosis of human disease. Dr. Pamela Ward directs the daily operation of this lab, which is located on the School of Medicine campus in Irvine. Along with implementing our clinical mission, this lab is involved in a variety of translational research projects with a broad aim of identifying, and/or characterizing the clinical relevance of molecular alterations in a variety of neoplastic, infectious and genetic diseases. Projects in this lab may be of interest to undergraduate and medical students, as well as Residents in the post-graduate Pathology training program (these projects are generally not appropriate for graduate students).
20.) Yan, H., Krishnan, K., Greenlund, A.C., Gupta, S., Lim, J.T.E., Schreiber, R.D., Schindler, C.W., and Krolewski, J.J. (1996) Phosphorylated interferon-alpha receptor 1 subunit (IFNaR1) acts as a docking site for the latent form of the 113kD STAT2 protein. EMBO J. 15:1064-1074.
22.) Yan, H., Krishnan, K., Lim, J.T.E., Contillo, L.G., and Krolewski, J.J. (1996) Molecular characterization of an interferon-alpha receptor subunit 1 (IFNaR1) domain required for tyk2 binding and signal transduction. Mol. Cell. Biol. 16:2074-2082.
33.) Krishnan, K., Singh, B., and Krolewski, J.J. (1998) Identification of amino acid residues critical for the Src-homology 2 domain-dependent docking of STAT2 to the interferon alpha receptor. J. Biol. Chem. 273:19495-19501.
39.) Nguyen, V.P., Saleh, A.Z.M., Arch, A.E., Yan, H., Piazza, F., Kim, J. and Krolewski, J.J. (2002) Stat2 binding to the interferon alpha receptor 2 (IFNaR2) subunit is not required for interferon-alpha signaling. J. Biol. Chem. 277:9713-9721.
40.) Saleh, A.Z.M., Nguyen, V.P. and Krolewski, J.J. (2002) Affinity of Stat2 for the subunits of the interferon-alpha receptor. Biochemistry. 41:11261-11268.
41.) Nastiuk, K.L., Kim, J. and Krolewski, J.J. (2003) Reduction of the mRNA encoding FLICE-like inhibitory protein during castration induced regression of the rat prostate. J. Cell Physiol. 196:386-393.
42.) Saleh, A.Z.M., Arch, A.E., Neupane, D., and Krolewski, J.J. (2004) Regulated proteolysis of the IFNaR2 subunit of the interferon-alpha receptor. Oncogene. 23:7076-7086.
43.) Kumar, K.G.S., Krolewski, J.J. and Fuchs, S.Y. (2004) Phosphorylation and specific ubiquitin-acceptor sites are required for ubiquitination and degradation of the IFNAR1 subunit of type I interferon receptor. J. Biol. Chem. 279:46614-46620.
44.) El Fiky, A., Arch, A.E. and Krolewski, J.J. (2005)
The intracellular domain of the IFNaR2 interferon receptor subunit
mediates transcription via Stat2. J. Cell. Physiol.
204:567-573.
45.) Krolewski, J.J. (2005) PROSPECTS: Cytokine and growth factor
receptors in the nucleus: what's up with that? J. Cellular
Biochem.
95:478-487.
46.) Saleh, A.Z.M., Greenman, K.L., Nguyen, D., Ehsan, P., Van
Vranken, D.L. and Krolewski, J.J. (2005) Madindoline A
binding to the extracellular domain of gp130. Biochemistry 44:10822-10827.
47.) Nastiuk, K.L., Liu, M.H., Hamamura, M., Muftuler, T., Nalcioglu,
O. and Krolewski, J.J. (2007) In vivo MRI volumetric measurement
of prostate regression and growth in mice. BMC Urology. 7:12.
48.) Nastiuk, K.L., Lo, K., Su, K. and Krolewski, J.J. (2007)
FLIP blocks TGF-ß1-induced caspase activation and apoptosis
in prostate epithelial cells. Mol. Cancer Res. 6:231-242.
49.) Cornforth, A.C., Davis, J., Khanifar, E., Nastiuk, K.L. and Krolewski, J.J. (2008) FOXO3a mediates the androgen-dependent regulation of FLIP and contributes to TRAIL-induced apoptosis of LNCaP cells. Oncogene IN PRESS. [view manuscript pdf]
50.) El Fiky, A., Pioli, P., Azam, A., Yoo, K., Nastiuk, K.L. and Krolewski, J.J. (2008) Nuclear transit of the intracellular domain of the interferon receptor subunit IFNaR2 requires Stat2 and Irf9. Cell. Signaling IN PRESS. [view manuscript pdf]
Manuscripts under review/revision:
51.) Nguyen, V.P., Zhang, H., Kadakia, S. and Krolewski,
J.J. Attenuation of interferon-alpha signaling by SOCS-1
and SOCS-3. [view manuscript pdf]
52.) Saleh, A.Z.M., Nastiuk, K.L., Truong, L., Black, R. and Krolewski, J.J. (2006) The interferon receptor IFNaR2 is cleaved
by TACE. [view manuscript
pdf]
53.) Yoo, K., Nastiuk, K.L. and Krolewski,
J.J. (2008) Transforming growth factor-ß1 induces apoptosis by suppressing FLICE-like inhibitory protein in DU-145 prostate epithelial cells. SUBMITTED. [view
manuscript pdf]
