Edward W. Harhaj, Ph.D.

Assistant Professor of Microbiology and Immunology
Room 503, Papanicolaou Building
1550 NW 10th Avenue
Telephone: 305-243-7893
Fax: 305-243-6410
Email: eharhaj@med.miami.edu

Research Interests:

Negative regulation of NF-kB and inflammation

The NF-kB transcription factor is critical for the induction of genes regulating innate and adaptive immunity.  NF-kB is tightly regulated to ensure that activation is only transient upon stimulation by cytokines such as tumor necrosis factor alpha (TNF-a) and interleukin-1 (IL-1).  Dysregulation of NF-kB leading to its persistent activation has been linked to chronic inflammation, autoimmunity and malignancy.  The zinc finger protein A20 (also known as TNFAIP3) has been shown to be essential for the termination of NF-kB signaling in tumor necrosis factor receptor (TNFR) and Toll-like receptor 4 (TLR4)/IL-1R signaling pathways by targeting key signaling molecules such as RIP1 and TRAF6 for inactivation.  Mice lacking A20 die prematurely of multi-organ inflammation and cachexia.  In addition, specific polymorphisms within the A20 genomic region have been associated with a number of human autoimmune diseases such as rheumatoid arthritis, systemic lupus erythomatosus and Crohn’s disease.  Additional studies have demonstrated loss-of-function mutations or deletions within A20 that are associated with lymphomas.  Despite the importance of A20 in limiting inflammatory responses, little is known about how A20 is regulated and also if other molecules are essential for the termination of NF-kB signaling.  Our laboratory has identified several key regulatory proteins that are required for the function of A20 and downregulation of NF-kB signaling.  We have found that TAX1BP1, Itch and RNF11 function in a nonredundant manner together with A20 to terminate NF-kB signaling.  In addition, the HTLV-I Tax oncoprotein promotes persistent NF-kB signaling and oncogenesis by targeting TAX1BP1 and inactivating the function of A20.  In future studies, we will determine the exact roles of each of the subunits of the A20 ubiquitin-editing complex by performing biochemical studies as well as gene targeting in mice.

Mechanisms of HTLV-I Tax-mediated NF-kB activation

Human T cell leukemia virus type I (HTLV-I) is a retrovirus that has been linked to the development of two distinct diseases in humans: 1) a neuroinflammatory disorder termed HTLV-I Associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP) that is clinically similar to multiple sclerosis (MS), and 2) a CD4+CD25+ malignancy known as adult T cell leukemia/lymphoma (ATLL).  NF-kB is one of the main targets of the HTLV-I encoded oncoprotein Tax to transform cells.  Furthermore, NF-kB is required for the survival of HTLV-I transformed cell lines and ATL leukemic cells.  Therefore, we are interested in determining the mechanisms of Tax-mediated NF-kB activation.  We have demonstrated that Tax promotes the relocalization of the IkB kinase (IKK) complex to the Golgi apparatus in a ubiquitin-dependent manner.  Targeting of IKK subunits to the Golgi is an essential step in Tax-mediated NF-kB activation, and we hypothesize that Tax utilizes the Golgi as an assembly platform for the generation of activated IKK subunits.  We have also found that Tax is polyubiquitinated in a nontraditional manner whereby the ubiquitin linkages are mediated through lysine 63 (K63) rather than lysine 48 (K48) linkages that promote proteosomal degradation.  We have identified the E2 ubiquitin conjugating enzyme Ubc13 as essential for both Tax ubiquitination and NF-kB activation.  Future studies will examine the requirements for Tax ubiquitination and other host factors that play a role in Tax-mediated NF-kB activation.

Selected Publications:

Shembade, N, K. Parvatiyar, N.S. Harhaj and E.W. Harhaj. 2009.  The ubiquitin-editing enzyme A20 requires RNF11 to downregulate NF-kB signaling. EMBO J. 28: 513-22.  [Related “Have you seen…?” article published in EMBO J. 28: 455-6].
 
Shembade, N, N.S. Harhaj, K. Parvatiyar, N.G. Copeland, N.A. Jenkins, L.M. Matesic and E.W. Harhaj. 2008.  The E3 ligase Itch negatively regulates inflammatory signaling pathways by controlling the function of the ubiquitin editing enzyme A20.  Nat. Immunol. 9: 254-262. [Related News and Views published in Nat. Immunol 9: 227-9; News and Commentary published in Immunol. Cell Biol. 86: 299-300; Faculty of 1000 citation: “Recommended”]

Shembade, N, N.S. Harhaj, M. Yamamoto, S. Akira and E.W. Harhaj.  2007. The HTLV-I Tax oncoprotein requires the ubiquitin conjugating enzyme Ubc13 for NF-kB activation.  J. Virol. 81: 13735-42.

Shembade, N, N.S. Harhaj, D.J. Liebl and E.W. Harhaj. 2007. Essential role for TAX1BP1 in the termination of TNF-a, IL-1 and LPS-mediated NF-kB and JNK signaling. EMBO J. 26: 3910-22. [Highlighted in Nature Immunology 2007 8: 1039].

Harhaj, N.S., B. Janic, J.C. Ramos, W.J. Harrington and E.W. Harhaj.  2007.  Deregulated expression of CD40 Ligand in HTLV-I infection: distinct mechanisms of downregulation in HTLV-I transformed cell lines and ATL patients.  Virology. 362: 99-108.

Harhaj, N.S., S.C. Sun and E.W. Harhaj.  2007.  Activation of NF-kB by the human T cell leukemia virus type I (HTLV-I) Tax oncoprotein is associated with ubiquitin-dependent relocalization of IKK.  J. Biol. Chem. 282: 4185-92.

Ramos, J.C., P. Ruiz, Jr., L. Ratner, I.M. Reis, C. Brites, C. Pedreso, G.E. Byrne, Jr., N. K. Toomey, V. Andela, E.W. Harhaj, I. Lossos and W.J. Harrington. 2007.  IRF-4 and c-rel expression in antiviral therapy resistant adult T-cell leukemia/lymphoma. Blood.  109: 3060-8.

Kamanaka, M., S.T. Kim, Y.Y. Wan, F.S. Sutterwala, M. Lara-Tejero, J.E. Galan, E. Harhaj and R.A. Flavell. 2006.  Expression of Interleukin-10 in intestinal lymphocytes detected by an Interleukin-10 reporter knock-in tiger mouse.  Immunity.  25: 941-52.

Harhaj, E.W., N.S. Harhaj, C. Grant, K. Mostoller, T. Alefantis, S.C. Sun and B. Wigdahl. 2005.  Human T cell leukemia virus type I Tax activates CD40 gene expression via the NF-kappa B pathway.  Virology.  333: 145-158.

Liao, G., M. Zhang, E.W. Harhaj and S.-C. Sun. 2004. Regulation of the NF-kB inducing kinase by TRAF3-induced degradation.  J. Biol. Chem.  279: 26243-50.

Xiao, G., M. Cvijic, A. Fong, E.W. Harhaj, M.T. Uhlik, M. Waterfield and S.-C. Sun. (2001)  Retroviral oncoprotein Tax induces processing of NF-kB2/p100 in T cells: evidence for the involvement of IKKa.  EMBO J.  20: 6805-6815.

Xiao, G.*, E.W. Harhaj* and S.-C. Sun. (2001) NF-kB-Inducing kinase regulates the processing of NF-kB2 p100.  Mol. Cell 7: 401-409.   *equal contribution.