L-selectin mediates the initial adhesion of lymphocytes to high endothelial venules (HEV) in lymph nodes during the process of lymphocyte homing. It also functions in leukocyte-endothelial interactions underlying the trafficking of leukocytes into chronic inflammatory sites. L-selectin functions as a lectin-like receptor by recognizing a discrete set of HEV-expressed ligands including GlyCAM-1, CD34, and podocalyxin. These ligands bear O-linked carbohydrate chains that are sulfated, fucosylated and sialylated. All three of these modifications are required for optimal recognition by L-selectin. A detailed analysis of GlyCAM-1 and CD34 has revealed that tile recognition determinant for L-selectin binding is a sulfated structure known as 6-sulfo sLex, a tetrasaccharide that possesses a sulfate ester on the C-6 position of N-acetylglucosamine. In the search for the sulfotransferase that elaborates this critical modification within HEV, the Rosen laboratory has cloned a family of GlcNAc-6-O-sulfotransferases. Two of these, known as GST-2 and GST-3, are present in HEV. GST-3 has been given the name HEC-GlcNAc6ST because of its highly restricted expression in high endothelial cells (HEC) of HEV. The direct involvement of HEC-GlcNAc6ST in elaborating L-selectin ligands has been established by disrupting this gene in mice. HEC-GIcNAc6ST knockout mice exhibit a significant but incomplete loss of HEV-expressed ligands for L-selectin and an impairment of lymphocyte homing to lymph nodes. The present grant will continue the study of HEC-GlcNAc6ST and the related enzyme, GST-2, with respect to their functions in lymphocyte homing and inflammatory leukocyte trafficking.
The specific aims are: 1) To determine the contribution of HEC-GIcNAc6ST to the activity of L-selectin ligands generated in situ; 2) To determine the expression of HEC-GlcNAc6ST in activated endothelium at sites of inflammation; 3) To determine the contribution of HEC-GlcNAc6ST to leukocyte recruitment and disease in mouse models of chronic inflammation; and 4) To determine the contribution of GST-2 to the generation of L-selectin ligands. Gaining further understanding of these sulfotransferases has considerable biomedical relevance, because these enzymes are potential therapeutic targets for blocking inflammatory diseases.
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