The chondrodystrophy, nanomelia: biosynthesis and processing of the defective aggrecan precursor.

The lethal chicken mutation nanomelia leads to severe skeletal defects because of a deficiency of aggrecan, which is the largest aggregating chondroitin sulphate proteoglycan of cartilage. In previous work, we have demonstrated that nanomelic chondrocytes produce a truncated aggrecan precursor that fails to be secreted, and is apparently arrested in the endoplasmic reticulum (ER). In this study, we investigated the biosynthesis and extent of processing of the abnormal aggrecan precursor. The truncated precursor was translated directly in cell-free reactions, indicating that it does not arise post-translationally. Further studies addressed the processing capabilities of the defective precursor. We found that the mutant precursor was modified by N-linked, mannose-rich oligosaccharides and by the addition of xylose, but was not further processed; this is consistent with the conclusion that it moves no further along the secretory pathway than the ER. Using brefeldin A we demonstrated that the defective precursor can function as a substrate for Golgi-mediated glycosaminoglycan chains, but does not do so in the nanomelic chondrocyte because it fails to be translocated to the appropriate membrane compartment. These studies illustrate how combined cell biological/biochemical and molecular investigations may contribute to our understanding of the biological consequences and molecular basis of genetic diseases, particularly those involving errors in large, highly modified molecules such as proteoglycans.