The San Diego Zoo's Conservation and Research for Endangered Species: Projects

Genome Studies for Conservation of the California Condor

Evaluating candidate loci for heritable chondrodystrophy in California condors

Our initial efforts focus on two genetic systems. The first of these is AGC1 (aggrecan 1; chondroitin sulfate proteoglycan 1). Aggrecan is one of the protoglyclans that form an integral part of the extracellular matrix in cartilaginous tissues, and is thought to have a role in cartilage’s ability to withstand compression. Mutations in this gene are involved in skeletal dysplasia in other species, such as chicken, human and mouse. Nanomelia, a hereditable syndrome in the chicken with phenotypic similarities to chondrodystrophy in condors, has been attributed to a truncated aggrecan protein, the result of a mutation in the AGC1 gene. Similar disorders were also described in turkey and Japanese quail but only in turkey it has been so far established that abnormal proteoglycan synthesis is involved in the disease manifestation (Dannenberg et al., 1982).

The AGC1 gene has similar 19-exon structure in birds and mammals, with the largest exon 12 encoding chondroitin sulphate attachment domain (CS). The latter has two subdomains, CS1 and CS2, and a variable repeat region (VNTR). The chicken VNTR includes 19 repeats of 60 nt (with the reported nanomelic stop codon), the human one has 19 repeats of 57 nt, and the bovine one 21 repeats of 57-63 nt. Repeat motif sequences vary between repeats within species as well as between species. The mouse and rat VNTRs have different repeat patterns, although a recognizable variant of the human repeat region can be seen in all species. So far, it was also shown in humans that number of repeats is really variable (13-33 repeats) but this could also be found in other species.

A human exon 12 single-base-par insertion introduced a frameshift of 212 aa
followed by a premature stop codon and resulted in a form of spondyloepiphyseal
dysplasia (SEDK). The appropriate paper by Gleghorn et al. (2005) described this
disease in a family that includes heterozygotes only. The mutation effect in
homozygotes was unknown, although the disease inheritance was demonstrated
to be autosomal dominant.

In the mouse, there are two reported Agc1 mutations. The first one is 7-bp deletion
in exon 5 that results in a frameshift and premature termination of the aggrecan
core protein. This causes the lethal embryonic cartilage matrix deficiency (cmd) in
homozygotes and postnatal developmental abnormalities and, eventually, death in
heterozygotes. The other mutation (cmd-Bc) is a near-complete deletion of the
whole aggrecan gene (exons 2 through 18) and is similar by the phenotype to

Thus, we may expect at least four possible scenarios of AGC1 mutations in

Gleghorn et al. (2005): Structure of the human AGC1 gene. A, Genomic organization of the AGC1 gene. Introns are represented by horizontal lines, exons are shown as vertical lines. Intron and exon sizes are shown in kb. B, cDNA structure of the AGC1 gene, showing the location of the exons. Exon sizes are shown in bp. The translation start and stop codons are labeled with ATG and TGA, respectively. The untranslated regions are shaded. The asterisk (*) marks the single-nucleotide (C) insertion at nucleotide 4015. C, Structure of the wild-type AGC1 protein. The location of each domain is marked by the amino acid numbers, with the initiation Met as amino acid 1. CS1 and CS2 p chondroitin sulfate attachment domains 1 and 2, respectively; G1, G2, and G3 p globular domains 1, 2, and 3, respectively; IGD p interglobular domain; KS p keratan sulfate attachment domain. The position of the insertion at amino acid 1214 is indicated by an asterisk (*). D, Structure of the mutant AGC1 protein identified in the SEDK-affected family. The 212-aa missense addition at the C-terminal is represented by a wave pattern. © 2005 by The American Society of Human Genetics.

Additional candidate genes for chondrodystrophy include RUNX2 (runt-related transcription factor 2) and Ihh [Indian hedgehog homolog (Drosophila)], both vital to the development of the vertebrate skeleton. Endochondral ossification, the process by which most bones are constructed, occurs when a cartilaginous template is gradually replaced by bony tissue. In early skeletal development, mesenchymal cells condense differentiate into chondrocytes. As chondrocytes multiply and begin to mature within the cartilaginous skeleton, they express Ihh. RUNX2 is also involved in chondrocyte proliferation and regulates Ihh expression. The limbs of Ihh -/- mice are severely diminished due to reduced chondrocyte proliferation.

We have identified in our BAC clone library the large pieces of DNA that contain the AGC1 and RUNX2 loci and have arranged to have the corresponding selected BAC clones sequenced in collaboration with Eric Green at the National Human Genome Research Institute (NHGRI). As a result, three positive BAC nucleotide sequences were obtained at the NIH Intramural Sequencing Center and deposited in GenBank (Accession Numbers AC171379, AC171743, and AC172166). (The value of this sequencing effort falls in the range of $45,000-$90,000, depending on the size of the inserts in the BAC clones.)

Obtaining the DNA sequences for these candidate genes will enable us to identify whether altered proteins are produced in normal and affected chicks, thereby explaining the effect of the mutation (as has been done for other genetic maladies including sickle cell anemia, cystic fibrosis, and Tay-Sachs disease).



Rationale for Sequencing Selected BAC Clones of the Genome of the California Condor (Gymnogyps californianus)
California Condor Genome Project: Introduction

Construction of a physical and genetic map of the California condor
California Condor Genetic Studies: Sex Determination, Identification of Clan Structure and Coping with a Genetic Disease

Back to Avian Genetics and Genomics

Wild Animal Park: Condor Ridge
California Condor Recovery Program
Milestones in California Condor Conservation

California Condor Mortality Challenges
Social Development and Reintroduction of California Condors
Studies of Courtship and Parental Care in Reintroduced California Condors in the U.S. and Mexico