Fig. 3

Ectopic mineralization phenotypes in dgf mutants. (A) Skeletal staining of a sibling, a dgf mutant, and a dgf mutant rescued by an enpp1 BAC transgene. Dgf mutants exhibit axial hypermineralization, resulting in partially fused vertebral centra. (B) Hypermineralization also affects the cleithra, which in dgf mutants typically exhibit nodule-like protrusions (arrow). no, notochord. (C) Meiotic map of the dgf locus. Recombinants per total number of mutants tested for each polymorphic marker are depicted in red, and markers used for mapping are in black. (D) Sequencing of dgf^hu4581 revealed a mutation in the splice acceptor site before exon 11 of the enpp1 gene. (E) The consequence at the transcript level is a deletion of 5 bp (Upper), resulting in a translational frameshift and a predicted stop codon after 23 amino acids (Lower). (F) Schematic representation of the BAC construct generated for the transgenic rescue of the dgf⁴⁵⁸¹ phenotype. Two genes are contained on the BAC, and kcnk3 was inactivated through a recombineering approach. (G) Dgf^sa156 fails to complement dgf^hu4581. (H) Schematic drawing of the predicted protein forms of enpp1, dgf^hu4581, and dgf^sa156, respectively. Enpp1 is a type II transmembrane protein. Striped box, transmembrane domain; light gray, somatomedin B-like binding domains; black, catalytic domain; dark gray, nuclease-like domain (36). See also Fig. S2.