Chondroitin Sulfate and Chondroitin/Keratan Sulfate Proteoglycans of Nervous Tissue: Developmental Changes of Neurocan and Phosphacan

Abstract

Abstract: We have studied developmental changes in the structure and concentration of the hyaluronic acid‐binding proteoglycan, neurocan, and of phosphacan, another major chondroitin sulfate proteoglycan of nervous tissue that represents the extracellular domain of a receptor‐type protein tyrosine phosphatase. A new monoclonal antibody (designated 1F6), which recognizes an epitope in the N‐terminal portion of neurocan, has been used for the isolation of proteolytic processing fragments that occur together with link protein in a complex with hyaluronic acid. Both link protein and two of the neurocan fragments were identified by amino acid sequencing. The N‐terminal fragments of neurocan are also recognized by monoclonal antibodies (5C4, 8A4, and 3B1) to epitopes in the G1 and G2 domains of aggrecan and/or in the hyaluronic acid‐binding domain of link protein. The presence in brain of these N‐terminal fragments is consistent with the developmentally regulated appearance of the C‐terminal half of neurocan, which we described previously. We have also used a slot‐blot radioimmunoassay to determine the concentrations of neurocan and phosphacan in developing brain. The levels of both proteoglycans increased rapidly during early brain development, but whereas neurocan reached a peak at approximately postnatal day 4 and then declined to below embryonic levels in adult brain, the concentration of phosphacan remained essentially unchanged after postnatal day 12. Keratan sulfate on phosphacan‐KS (a glycoform that contains both chondroitin sulfate and keratan sulfate chains) was not detectable until just before birth, and its peak concentration (at 3 weeks postnatal) was reached ∼1 week later than that of the phosphacan core protein. Immunocytochemical studies using monoclonal antibodies to keratan sulfate (3H1 and 5D4) together with specific glycosidases (endo‐β‐galactosidase, keratanase, and keratanase II) also showed that with the exception of some very localized areas, keratan sulfate is generally not present in the embryonic rat CNS.