University of Virginia Health System

RESEARCH:

Our laboratory is interested in Ras signal transduction and the molecular mechanisms of tumor suppression, specifically within the novel family of tumor suppressors represented by NORE1 and RASSF1.

In humans, Ras is the most common dominant oncogene, especially in tumors of epithelial origin. Ras activation often leads to uncontrolled cell proliferation and the death of the organism. Therefore, it is likely that there are molecular mechanisms in cells that sense the suitability of Ras activation, and in the case of inappropriate activation, eliminate the cell. One of the mechanisms used to eliminate unwanted or potentially dangerous cells is apoptosis. NORE1 was discovered as a protein capable of binding to activated Ras in vitro and in vivo with high affinity.  We found that NORE1 could bind to Ras in transfected cells and initiate an apoptotic response through a Ser/Thr protein kinase, MST1. MST1 appears to be constitutively bound to NORE1 and this complex could bind Ras upon cell stimulation.  In transfected cells, the Kirsten isoform of the Ras oncogene product is capable of inducing apoptosis -- which is NORE1 and MST-dependent.  Recent data from several laboratories strongly suggests that NORE1, specifically its longest splice isoform NORE1A, is a tumor suppressor.  We hypothesize that NORE1 is the key regulator in Ras activation, determining an outcome of proliferation or apoptosis, shifting the balance towards the latter upon binding Ras. To understand the molecular mechanisms of NORE1 tumor suppression, we are planning to identify Ras isoform(s) or Ras-like GTPases that bind NORE1 in vivo and physiological events that influence this binding. We also plan to reveal molecular mechanisms by which the association of NORE1, with its small GTPase partner, results in tumor suppression.

RASSF1 is located on human chromosome 3p21, among a cluster of putative tumor suppressor genes.  This cluster is frequently deleted in lung tumors, as well as in many breast, renal, hepatic, prostate, head/neck and other cancers.  The RASSF1 gene is expressed in multiple splice forms, each containing a C-terminal Ras Association domain with alternative N-terminal segments.  The expression of the longer RASSF1A isoform is selectively extinguished by promoter methylation in many human tumors, whereas the shorter RASSF1C splice variant continues to be expressed.  Moreover, forced expression of the RASSF1A polypeptide in such tumor cell lines substantially suppresses their growth rate in vitro and in vivo. RASSF1 is closely related to NORE1; however, RASSF1 cannot bind Ras-GTP.  Nevertheless, RASSF1 proteins can bind the proapoptotic kinase MST1.

To understand the molecular mechanisms of RASSF1A tumor suppression, we are going to investigate what activates RASSF1A and determine its downstream effectors.  We will also explore the role of MST kinases in RASSF1A tumor suppression.

REFERENCES:

• Moshnikova A., Frye J., Shay J. W., Minna J. D., and Khokhlatchev A.V. The growth and tumor suppressor NORE1A is a cytoskeletal protein that suppresses growth by inhibition of the ERK pathway. J. Biol. Chem., 2006, in press (accepted on January 18, 2006).
  
• Praskova M., Khoklatchev A., Ortiz-Vega S., Avruch J. Regulation of MST1 kinase by autophosphorylation, by the growth inhibitory proteins, RASSF1 and NORE1 and by Ras. Biochem J., 2004, Jul 15; 381(Pt 2):453-462.
• Rabizadeh S., Xavier R. J., Ishiguro K., Bernabeortiz J., Lopez-Ilasaca M., Khokhlatchev A., Mollahan P., Pfeifer G. P., Avruch J., Seed B.  The scaffold protein CNK1 interacts with the tumor suppressor RASSF1A and augments RASSF1A-induced cell death.  J. Biol. Chem., 2004, Jul 9; 279(28): 29247-29254.
  
• Khokhlatchev, A., Xavier, R., Rabizadeh, S., Nedwidek, M., Chen, T., Zhang, X,-f., Seed, B., Avruch, J. Identification of a novel Ras-regulated proapoptotic pathway. Current Biology, (2002) 12 (4) 253-265.
• Ortiz-Vega, S., Khokhlatchev, A., Nedwidek, M., Zhang, X,-f., Dammann, R., Pfeifer, G.P., Avruch, J. The Putative Tumor Suppressor RASSF1A Homodimerizes and Heterodimerizes with the Ras-GTP Binding Protein Nore1. Oncogene, (2002) 21(9) 1381-1390.
• Lin, Y., Khokhlatchev, A., Figeys, D., Avruch, J. Death-associated protein 4 binds MST1 and augments MST1-induced apoptosis J. Biol. Chem. (2002) 277 (50) 47991-48001.