Correlation between developmental chondroitin sulfate pattern and the upregulation of chondroitin 6-sulfotransferase mRNA expression in the embryonic hindbrain

Abstract

Reports of chondroitin sulfate (CS) glycoforms in boundary regions of mouse embryonic hindbrain led us to hypothesize that these glycoforms are involved in patterning hindbrain segmentation. Immunohistochemical study using monoclonal antibody CS-56 on E 8.5 to E 11.5, the time when hindbrain segmentation occurs, revealed immunopositivity in the neuroepithelial boundaries in both tissue sections and whole-mount embryos. The signals in E8.5 and E 9.5 embryos were found stronger than those in later stages. Competitive neutralization of CS-56 immunoreactivity with different CSs found 6-sulfated glycoforms superior to other CS glycoforms, suggesting the abundance of 6-sulfated CSs. The result was confirmed by analyzing the chondroitinase digestion products of the different CS glycoforms using fluorophore-assisted carbohydrate electrophoresis (FACE). As C6S is a product of chondroitin 6-sulfotransferase (C6ST)-catalyzed reaction, upregulation of C6ST gene expression in the boundary regions was anticipated. Riboprobes specific for the C6ST mRNA were synthesized and used for in situ hybridization. The anti-sense probe indicated distribution of positive cell bodies in the hindbrain neuroepithelium of E8.5 and E9.5 embryos but not in the later stages. We have therefore found temporal correlation between the expression of C6ST mRNA and the product of C6ST activity. The basis for the spatial difference in expression patterns of gene activity and enzyme activity remains to be investigated. (SPON: The Hong Kong Society of Neurosciences). Supported by RGC Grant # HKU7294/01M