Role of the Raf-1, B-Raf, and MEK-1 kinases in apoptosis

The cytosolic serinehreonine kinases Raf and Mek play a central role in evolutionary conserved signal transduction pathways. These enzymes have been implicated in a variety of biological processes, such as regulated and deregulated proliferation, differentiation, and programmed cell death (apoptosis). Because of this, they are considered attractive therapeutic targets. Gene ablation experiments conducted by us (Raf-1 and Mek-1) and by others (B-Raf) have revealed that Raf-1 and B-Raf exert essential anti-apoptotic functions that cannot be rescued by the other Raf isoforms. Surprisingly, ablation of their common effector MEK-1 confers resistance towards matrix detachment-induced apoptosis. Thus, these kinases must target different steps of the apoptotic process. The aim of this project is to identify the molecular mechanism(s) underlying the unique functions of Raf-1, B-Raf, and Mek-1 in apoptosis induced by Fas stimulation or by matrix detachment. To this end, we will use mouse strains and cell lines produced in the lab (conventional Raf-1 and Mek-1 knock-out, conditional Raf-1 knock-out) or obtained from collaborating groups (conditional B-Raf). We will combine phenotype analysis in the context of the whole organism (in vivo) as well as of cells (ex vivo) with biochemical experiments in ablated cells to elucidate the molecular basis of a given phenotype. The results of these experiments will help defining the function(s) of Raf-1, B-Raf and Mek-1 and their relevant biological targets. The information obtained will be instrumental in assessing the potential of these molecules as therapeutic targets and in directing the design and use of pharmacological kinase inhibitors.