EXPRESSION OF THE CARTILAGE MATRIX PROTEIN GENE AT DIFFERENT CHONDROCYTE DEVELOPMENTAL STAGES

Cartilage matrix protein (CMP), a major noncollagenous component of ce rtain types of hyaline cartilage, is synthesized by chondrocytes in a developmentally regulated manner, In this study, we monitored the accu mulation of CMP in the developing chicken limb and sternum by immunost aining. In older embryos, the specific extracellular staining was rest ricted to the resting/proliferative zone of metaphyseal cartilage and to the immediately adjacent hypertrophic cartilage. A lack of staining was observed in the peripheral layers of articular cartilage, Data we re compared with the accumulation of CMP mRNA measured by Northern ana lysis relative to other cartilage-specific messages in cell cultures r epresenting different stages of chondrocyte differentiation, as well a s with the steady state mRNA levels in tissue samples. We found a corr elation between the gene expression pattern of the in vitro cultures a nd the one observed in certain in vivo differentiation stages. The hig h-density mesenchyme culture was utilized as a model for studying the events at early stage I (stage Ia) of chondrogenesis. This culture was characterized by relatively low steady state mRNA levels for cartilag e proteins, including the later activation of the CMP gene as compared to type II collagen or link protein genes, and relatively high steady state mRNA levels for type VI collagen and beta-actin. Chicken embryo chondrocyte cultures obtained from sterna of 14-day-old embryos, howe ver, consisted predominantly of stage Ib chondrocytes, and showed high steady state levels for cartilage proteins, but relatively lower leve ls for type VI collagen and beta-actin mRNAs. In accordance with the i n vivo data, a relatively high steady state level was detected for CMP mRNA in cultures of hypertrophic (stage II) chondrocytes. We also per formed transient expression assays in the various culture systems to s tudy the role of the promoter upstream and intronic control regions in the tissue- and developmental stage-specific regulation of the CMP ge ne. We showed that the enhancer worked in a lineage specific manner, b y further stimulating the minimal promoter activity independent of the developmental stage of chondrocytes, while it did not in other tissue s. The promoter upstream control regions, however, seemed to play a ro le in restricting the promoter activity to a certain chondrocyte devel opmental stage.