Abstract
The distribution of heparan sulfate (HS) proteoglycans in clonal rat parathyroid cells is regulated by the extracellular Ca2+ concentration, which is a principal factor for parathyroid cell function (Takeuchi, Y., Sakaguchi, K., Yanagishita, M., Aurbach, G. D., and Hascall, V. C. (1990) J. Biol. Chem. 265, 13661-13668). Increasing the concentration of extracellular Ca2+ in the physiological range redistributes HS proteoglycans from the cell surface to an intracellular compartment. We have now examined effects of the extracellular Ca2+ concentration on the metabolism of the HS proteoglycans in detail using [35S]sulfate metabolic labeling-chase experiments. Two distinct metabolic pathways were demonstrated: (i) the intracellular generation of HS chains from HS proteoglycans in prelysosomal compartments followed by their release into the medium (pathway 1), and (ii) intracellular generation of HS oligosaccharides from HS chains in prelysosomal compartments, which are eventually degraded into free sulfate in lysosomes (pathway 2). The HS oligosaccharides were exclusively present within the cells, whereas HS chains were found primarily in the medium. The cells do not internalize either HS proteoglycans or HS chains from the medium. These observations indicate that these two degradation pathways are independent. In addition to these pathways, approximately 15% of the HS proteoglycans were released into the medium as a proteoglycan form. Treatment of cells with chloroquine, a lysosomotropic agent, did not affect generation of HS chains but inhibited conversion of HS chains to HS oligosaccharides or to free sulfate and resulted in the release of HS chains from the cells. The drug did not affect metabolic pathway 1. The extracellular Ca2+ concentration did not alter these intracellular degradation pathways for HS proteoglycans in the parathyroid cells. Thus, extracellular Ca2+ appears to regulate only the distribution of HS proteoglycans between the cell surface and intracellular compartments, and the process of cycling between these compartments when extracellular Ca2+ is low.
Highlights
The distribution of heparan sulfate (HS) proteogly- distributed throughout animal tissues
We have examined effects of the extracellular Ca" concentration on the metabolism of the HS proteoglycans in detail using [3SSlsulfatemetabolic labeling-chase experiments
We have reported [6] that thecellular distribution of HS proteoprelysosomal compartments followed by their release glycans synthesized by this cell line is regulated by [Ca"], into the medium, and (ii) intracellular such that the proportion of HS proteoglycans on the cell generation ofHS oligosaccharides from HS chains in surface increases, while intracellular HS proteoglycans recipprelysosomal compartments, which are eventually de- rocally decrease as [Ca"], is reduced
Summary
14677-14684,1992 Printed in U.S.A. Metabolic Pathways of Heparan Sulfate Proteoglycans in a Rat Parathyroid Cell Line*. The cells donot internalize either HS similar to those for recycling receptors, such as transferrin proteoglycans or HS chains from the medium These receptors or low density lipoprotein receptors [7,8]. Another interesting featureof HS proteoglycans in the rat appears to regulate only the distributionof HS proteo- parathyroid cells is that intermediate products in intracellular glycans between the cell surface and intracellulacrom- degradation processes are not readily apparent This differs partments, and the process of cycling between these from degradation processes of HS proteoglycans observed in compartments when extracellular Ca2+is low. Cell surface heparan sulfate(HS)' proteoglycans are widely the metabolism of HS proteoglycans in parathyroid cells to define degradation pathways inmore detail. 0.15 M NaCI, 20 mM HEPES, pH 7.4, containing the same concentrations of Ca2+ and Mg" as the labeling medium
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