During development and maintenance of tissues there is a balance between cell division and cell death in order to generate the proper number and type of cells in the animal. This process includes the elimination by programmed cell death (PCD) or apoptosis of cells that are harmful or not longer needed. Genes that function as effectors or suppressors of the PCD pathway have been identified in Caenorhabditis elegans. Within mammalian systems, the bcl-2 proto-oncogene has been shown to serve as a suppressor of PCD. However, Bcl-2 failed to block several forms of PCD arguing that certain cell deaths are independent of Bcl-2. We have identified a novel gene, bcl-x, that functions as a dominant regulator of apoptotic cell death in vertebrates including humans. The coding region of bcl-x displays extensive amino acid homology with Bcl-2. Two forms of the Bcl-x protein, Bcl-x1 and Bcl-xs, are expressed in human cells by alternative splicing of the bcl-x mRNA. Bcl-x1 is a powerful repressor of PCD as demonstrated by gene transfer experiments. By contrast, Bcl-xs is a facilitator of cell death by inhibiting the death suppressor activity of Bcl-2 and perhaps Bcl-x1. Importantly, the expression of both bcl-x1 and bcl-xs mRNA is highly regulated in the resting state in tissues and during cell activation arguing that bcl-x plays an important role in determining cell fate in the animal. The current proposal will focus on the murine bcl-x to enable us to assess in detail its developmental regulation and biological function. The murine full-length cDNA has been isolated and sequenced. Genomic clones encompassing the bcl-x locus have been isolated. Furthermore, antibodies directed against the murine Bcl-x proteins and a panel of stable bcl-x transfectants have been developed.
The specific aims of the present application are to (i) define the genomic organization and establish the chromosomal localization of the mouse bcl-x gene (ii) examine the expression of Bcl-x1 and Bcl-xs in tissues and their subcellular location (iii) to identify interaction between Bcl-x proteins and cellular proteins including Bcl-2 and (iv) target bcl-x transgenes to B cells to examine their biological role in the animal. Beyond its critical role in regulating physiological cellular processes. genes that control the PCD pathway such as bcl-2 and bcl-x have been implicated in the pathogenesis of B-cell lymphoma and autoimmune disease arguing that deregulation of PCD genes plays a role in disease development. Therefore, further understanding of bcl-x should provide ways to manipulate the cell death mechanism and ultimately to develop novel therapeutic strategies for cancer, autoimmune, and degenerative diseases.
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