Insertional mutation in the Golgb1 gene is associated with osteochondrodysplasia and systemic edema in the OCD rat.

Homozygous rats (ocd/ocd) of a mutant inbred strain, OCD (osteochondrodysplasia), show osteochondrodysplasia, systemic edema, cleft palate, protruding tongue, disproportionate dwarfism, and lethality immediately after birth. Their epiphyses show decreased levels of glycosaminoglycans and weak staining for extracellular matrix proteins. The epiphyseal chondrocytes have large vesicles and expanded endoplasmic reticulum and Golgi apparatus. These phenotypic features are inherited in an autosomal recessive manner, and the ocd locus responsible for these phenotypes has been mapped close to D11Mgh3 on rat chromosome 11. In the present study, we characterized the embryonic pathogenesis of ocd/ocd rats and identified the mutant gene. Subcutaneous edema in the dorsal portion was found at embryonic day (E) 16.5, and the other anomalies described above were apparent after E18.5 in ocd/ocd. Whole mount immunohistochemistry for Sox9 revealed that mesenchymal condensation was delayed in limb bud in ocd/ocd, and skeletal preparation showed that the progression of whole-body chondrogenesis was delayed in ocd/ocd. Histological and immunohistological analyses of the femur showed that cell proliferations of resting and proliferative zones of growth plate were significantly reduced in ocd/ocd embryos. Fine linkage mapping localized the ocd locus within 84kb of positions 65,584-65,668kb containing a part of Golgb1 gene on chromosome 11. Expression of Golgb1 mRNA was found in limb buds, somite derivatives and calvaria. Sequence analysis identified a 10-bp insertion in exon 13 of the Golgb1 gene in ocd/ocd rats. The Golgb1 gene encodes the COPI vesicle tethering factor, giantin. This insertion mutation causes a frame shift, and introduces a premature termination codon at codon 1082, leading to truncation of the C-terminal two thirds of giantin. By in-gel Western analysis using anti-giantin antibody that recognizes an epitope within 200 aa of the C-terminus, the expression of giantin was not detected in ocd/ocd embryos. As the C-terminal region of giantin is required for localization to the Golgi apparatus, these results strongly suggested that giantin is functionally defective in ocd/ocd rats. Therefore, we concluded that mutation of the Golgb1 gene is responsible for the phenotypic characteristics including osteochondrodysplasia of ocd/ocd, and that giantin plays a pivotal role in multiple aspects of chondrogenesis.