The intracellular contents of eukaryotic cells are sub-divided into discrete compartments, membrane enclosed organelles, which provide unique environments for specific cellular functions. Under special circumstances, such as during terminal differentiation of red blood cells and lens fiber cells or in programmed cell death, organelles are degraded. This critical process must be tightly regulated to insure that activation of the degradation process occurs at specific times and is directed towards distinct target membranes. Nevertheless, little is known about these critical aspects of this important cellular process. In this proposal we describe a comprehensive study of the mechanism by which organellar degradation occurs. Our studies will focus on the mechanism by which organellar degradation occurs. Our studies will focus on the cell biology and biochemistry of this degradative process during maturation of red blood cells. In our preliminary studies we have shown that 15-lipoxygenase (15-LOX) has the capability to both trigger and participate in organellar degradation that occurs during differentiation of red blood cells, by insertion into and permeabilization of target cellular membranes. This is followed by release and subsequent degradation of organellar contents. We further propose to study the mechanism for this action of 15 LOX on specific membranes, to define and isolate cellular factors involved in this degradation process. The in vivo significance of such factors will be examined by over-expressing or micro-injection of 15-LOX in murine erythroleukemia cells, a model for cell differentiation. These studies will enable us to determine whether over-expression of this enzyme is sufficient for differentiation of red cells. In addition, we will inject rabbits with known inhibitors of 15- LOX to ascertain whether this treatment alters organellar degradation and subsequent red cell differentiation.
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