Chick cartilage fibronectin differs in structure from the fibronectin in limb mesenchyme

Fibronectin, present in the extracellular matrix of several tissues, is heterogeneous in structure. This heterogeneity is largely due to the alternative splicing of three exons (IIIB, IIIA, and V) during processing of the fibronectin primary transcript. Previously, we determined that the splicing patterns of fibronectin mRNAs change from B+A+ to B+A- during the differentiation of mesenchyme into cartilage (V. D. Bennett, K. M. Pallante, and S. L. Adams, J. Biol. Chem. 266, 5918-5924, 1991). Therefore, the structure of fibronectin at the protein level most likely changes during chondrogenesis as well. In order to characterize the fibronectin protein in chick limb prechondrogenic mesenchyme and cartilage, we generated a polyclonal antibody specific for the region encoded by exon IIIB in the chick fibronectin gene. Immunoblot and immunohistochemistry analyses with this antibody and an antibody specific for the region encoded by exon IIIA indicate that both antibodies react with the fibronectin present in prechondrogenic mesenchyme. In contrast, only the exon IIIB antibody reacts with the fibronectin present in chick cartilage. Quantitative ELISA assays with these antibodies indicate that approximately 96% of the fibronectin in chick embryonic cartilage contains exon IIIB while less than 3% of the fibronectin contains exon IIIA. These results corroborate the structures of the fibronectin isoforms present in prechondrogenic mesenchyme and cartilage that were predicted from our previous characterization of the fibronectin mRNAs in these tissues and indicate that essentially all of the fibronectin in cartilage is synthesized and secreted by cartilage chondrocytes.