Abstract
Heparanase is a mammalian endoglycosidase that degrades heparan sulfate (HS) at specific intrachain sites, an activity that is strongly implicated in cell dissemination associated with metastasis and inflammation. In addition to its structural role in extracellular matrix assembly and integrity, HS sequesters a multitude of polypeptides that reside in the extracellular matrix as a reservoir. A variety of growth factors, cytokines, chemokines, and enzymes can be released by heparanase activity and profoundly affect cell and tissue function. Thus, heparanase bioavailability, accessibility, and activity should be kept tightly regulated. We provide evidence that HS is not only a substrate for, but also a regulator of, heparanase. Addition of heparin or xylosides to cell cultures resulted in a pronounced accumulation of, heparanase in the culture medium, whereas sodium chlorate had no such effect. Moreover, cellular uptake of heparanase was markedly reduced in HS-deficient CHO-745 mutant cells, heparan sulfate proteoglycan-deficient HT-29 colon cancer cells, and heparinase-treated cells. We also studied the heparanase biosynthetic route and found that the half-life of the active enzyme is approximately 30 h. This and previous localization studies suggest that heparanase resides in the endosomal/lysosomal compartment for a relatively long period of time and is likely to play a role in the normal turnover of HS. Co-localization studies and cell fractionation following heparanase addition have identified syndecan family members as candidate molecules responsible for heparanase uptake, providing an efficient mechanism that limits extracellular accumulation and function of heparanase.
Highlights
Heparanase is a mammalian endoglycosidase that degrades heparan sulfate (HS) at specific intrachain sites, an activity that is strongly implicated in cell dissemination associated with metastasis and inflammation
These results suggest that heparanase, secreted by each of the cell lines, rapidly interacts with membranous heparan sulfate proteoglycans (HSPGs), which is presumably followed by uptake and internalization and is competed by the addition of heparin
Chlorate treatment had no such effect on extracellular heparanase accumulation (Fig. 1), suggesting that N- rather than O-sulfate groups mediate the interaction of HS with heparanase, as indicated by the inability of N-acetylated heparin to promote extracellular accumulation of heparanase (Fig. 1F)
Summary
Heparanase is a mammalian endoglycosidase that degrades heparan sulfate (HS) at specific intrachain sites, an activity that is strongly implicated in cell dissemination associated with metastasis and inflammation. Co-localization studies and cell fractionation following heparanase addition have identified syndecan family members as candidate molecules responsible for heparanase uptake, providing an efficient mechanism that limits extracellular accumulation and function of heparanase. Heparanase is a mammalian endo--D-glucuronidase that cleaves heparan sulfate (HS) side chains at a limited number of sites [1,2,3] Such enzymatic activity is thought to participate in degradation and remodeling of the extracellular matrix and to facilitate cell invasion associated with cancer metastasis and inflammation (1, 4 – 6). In addition to its intimate involvement in the egress of cells from the blood stream, heparanase activity may release a multitude of HS-bound, extracellular matrix-resident growth factors, cytokines, chemokines, and enzymes that might profoundly affect cell and tissue function [1, 21]. Co-localization and cell fractionation studies following heparanase addition have identified syndecans, rather than glypicans, as candidate HSPGs responsible for heparanase uptake
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