The ecto-enzyme 5'- nucleotidase (5'-NT, CD73) catalyzes the extracellular dephosphorylation of purine and pyrimidine ribo- and deoxyribonucleoside monophosphates to their corresponding nucleosides. 5'-NT expression is tightly regulated during lymphocyte differentiation, with 5'-NT activity increasing five to tenfold as T- cells and B-cells mature. This pattern of expression suggests an important function for 5'-NT in mature versus immature lymphocytes, but that function remains undefined. Curiously, in the thymus, the pattern of expression of adenosine deaminase (ADA), the next enzyme in the purine salvage pathway, is reciprocal to that of 5'- NT. The reason for the inverse relationship between these two enzymes which catalyze consecutive reactions in purine catabolism is not understood. The principal investigator proposes that 5'-NT, by catalyzing the production of adenosine from AMP, and ADA, by catalyzing the degradation of adenosine to inosine, function together to control the availability of extracellular adenosine for interaction with cell surface adenosine receptors. To gain additional insight into the function of 5'-NT during lymphocyte development and to obtain evidence for functional coupling of 5'-NT to adenosine receptor engagement, the investigators propose the first four specific aims below. They also propose to analyze mechanisms of 5'-NT gene regulation.
The Specific Aims are: I. To develop transgenic mice which express the human 5'-NT gene inappropriately at early stages of T-cell and B-cell development and examine the consequences of lymphocyte maturation. II. To develop a 5'-NT knock-out mouse and study its lymphocyte differentiation. III. To analyze adenosine receptor expression during lymphocyte development and differentiation. IV. To evaluate the ability of 5'-NT and ADA to control adenosine receptor engagement. V. To analyze 5'-NT gene regulation. The investigators believe the results of the proposed experiments will not only increase our knowledge of how the cell type and developmental stage specific patterns 5'-NT gene expression are controlled, but will also demonstrate the consequences of aberrantly regulated 5'-NT expression in cells of lymphoid linage. Adenosine receptors mediate a variety of physiologic responses and are potential targets for pharmacologic intervention. It is, therefore, important to understand the consequences of adenosine receptor engagement upon lymphocyte development and immune responses. The investigators believe that their results may suggest new strategies for modulation of immune system development or function via manipulation of 5'-NT expression and/or lymphocyte adenosine receptor engagement.
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