Cooper is an essential trace element with plays a fundamental role in the biochemistry of all aerobic organisms. The long-term objective of these studies is to define the role of copper in embryonic and fetal development. Although numerous experimental and clinical studies reveal an essential role for cooper in prenatal development the mechanisms underlying these observations are unknown. Recent studies reveal that the intracellular delivery of cooper to specific targets is determined by a family of proteins termed copper chaperones. Disruption of the genes encoding the murine copper chaperone atox1 results in growth retardation and skeletal anomalies offering the opportunity to examine the mechanisms of alterations in copper homeostasis on skeletal morphogenesis in a well-defined genetic model. The studies in this proposal are intended to elucidate the function of atox1 in chondrocyte copper homeostasis and to examine the effects of atox1-associated copper deficiency on the genetic programs determining bone growth and development. The molecular and cellular mechanisms of copper homeostasis in chrondrocytes will be studied by analyzing copper uptake, trafficking, efflux and incorporation into lysyl oxidase following 64Cu metabolic labeling of cells from atox1+/+ and atox1+/+ mice. Genes disrupted or with probes generated from atox1+/+ and atox1-/- chondrocytes isolated by laser capture microdissection. Finally, examined by characterization of the phenotype of atox1-/- mice bred onto the genetic background of deficiencies in fibroblast growth factor receptors and heparan sulfate proteoglycans known to be essential for skeletal morphogenesis. Taken together the results of these studies will permit insight into the mechanisms of cooper homeostasis in developing bone as well as a direct analysis of opposing influences of genetically determined nutritional influences on the expression of genes affecting fetal growth and morphogenesis.
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