The biochemically and immunologically distinct CSPG of notochord is a product of the aggrecan gene.
Using the monoclonal antibody S103L, which reacts specifically with an epitope in the chondroitin sulfate-rich domain of the chick cartilage chondroitin sulfate proteoglycan (CSPG) core protein, we have identified the predominant CSPG expressed by notochord. This large notochord CSPG is first detected immunohistochemically as early as stage 16, long before chondrogenesis occurs, and is expressed continuously during the time of active neural crest migration and through the onset of sclerotomal differentiation. Because of the cross-reactivity of both notochord and cartilage CSPGs with the S103L antibody, extensive molecular and biochemical analysis of the two CSPGs was carried out. Striking differences distinguish the notochord and cartilage (aggrecan) CSPGs at the level of posttranslational modification. Notably, cartilage aggrecan carries a significant content of keratan sulfate (KS) chains, while the notochord CSPG is devoid of KS. In contrast, cartilage aggrecan lacks the HNK-1 epitope, while the notochord CSPG has a high content of HNK-1. Three different approaches were used to establish the relationship of the two CSPGs at the molecular level. Northern blot analysis, using aggrecan probes, detected same-sized messages from notochord and cartilage RNA. Overlapping fragments, generated by RT-PCR using primers covering 98% of the entire coding sequence from the known cartilage structure, were of identical size in notochord and cartilage. Taking advantage of our recent studies, which demonstrated a single base change in the aggrecan gene resulting in conversion of Glu to a STOP codon in exon 12 of chick aggrecan as the molecular basis of the defect nanomelia, we demonstrated that the same mutation was present in notochord mRNA from nanomelic chicks. These results provide evidence that the chick aggrecan gene is expressed very early in development in notochord and confirm that the core proteins expressed in chick notochord and cartilage are derived from the same gene. These findings strongly support the hypothesis that the final structural characteristics of each proteoglycan are determined not only by the core protein but also by tissue-specific, developmentally regulated posttranslational mechanisms, functioning within the context of the requirement for specific extracellular matrices.
Domowicz
M
M
Department of Pediatrics, University of Chicago, Illinois 60637, USA.
Li
H
H
Hennig
A
A
Henry
J
J
Vertel
B M
BM
Schwartz
N B
NB
D053674:Aggrecans; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001483:Base Sequence; D002356:Cartilage / Q000737:chemistry; D002642:Chick Embryo; D011508:Chondroitin Sulfate Proteoglycans / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D016326:Extracellular Matrix Proteins; D005455:Fluorescent Antibody Technique; D037181:Lectins, C-Type; D051379:Mice; D008969:Molecular Sequence Data; D009672:Notochord / Q000737:chemistry*; D011509:Proteoglycans / Q000235:genetics*; D011817:Rabbits
C508320:Acan protein, mouse; D053674:Aggrecans; D000911:Antibodies, Monoclonal; D011508:Chondroitin Sulfate Proteoglycans; D016326:Extracellular Matrix Proteins; D037181:Lectins, C-Type; D011509:Proteoglycans