Significant research efforts have focused on the tumor suppressor proteins involved in inhibition of cell cycle progression and induction of programmed cell death in cancer models. Several lines of evidence have established a role for Ras association family (RASSF) member RASSF1A as a tumor suppressor protein. (1) RASSF1A has been shown to play a key role in tumor necrosis factor-1 (TNF1) induced programmed cell death or apoptosis. (2) In agreement with a proposed role for RASSF1A as a tumor suppressor protein, several studies have described epigenetic inactivation of the gene encoding RASSF1A in malignant cancers by methylation of the promoter for exon 1 of the RASSF1A gene. (3) There are several polymorphisms within the coding region of RASSF1A that are evident in several forms of cancer. (4) Finally mice that do not express RASSF1A develop spontaneous cancers at a higher rate than their wild type littermates. The majority of these studies have focused on the role of RASSF1A in many tumors and cell lines of epithelial origin, including lung, kidney and breast tissues. The possible role of this protein in lymphocyte signaling and apoptosis has been the subject of considerably less investigation, although analyses of methylation-induced promoter silencing of a wide range of tumors have revealed that RASSF1A is silenced in some forms of leukemia and lymphoma. The major long-term objective of this research proposal is to understand the role of RASSF1A in transcriptional regulation of genes that are involved in lymphocyte function and homeostasis. Our preliminary results have identified a role for RASSF1A in transcriptional regulation of a key lymphocyte apoptosis regulator, FasL, in the human T leukemia cell line Jurkat. The specific regulatory elements within the FasL promoter that are affected by RASSF1A expression will be identified through a series of cotransfection studies using elements of the FasL promoter as reporters. The contribution of specific functional domains of RASSF1A to transcriptional activation and to T cell signaling pathways, including calcium flux and phosphorylation, will be analyzed by expression of mutant forms of RASSF1A and pharmacological modulators of calcium level and PTK activity.
This proposal seeks to define the specific molecular interactions of RASSF1A with cellular components that collaborate in T cell apoptosis. RASSF1A is silenced by methylation-induced inactivation in many tumors, which make it a potential target of demethylating reagents that are widely used in clinical trials for cancer treatment. Understanding the role of RASSF1A in cell function is critical to predict possible outcomes of expression/re-expression of this protein.
