Duke University
Project 2
Title: Mechanism of Stress-induced Developmental Abnormalities
PI: Jonathan Freedman, PH.D.
Exposure to transition metals (cadmium, copper) and organic chemical (PCB's, endosulfan) disrupts normal cellular development and differentiation. These hazardous chemicals have been shown to induce intracellular oxidative stress and produce reactive oxygen species (ROS). ROS in turn can stimulate activating protein-1 (AP-1) binding, as well as increase c-jun and c-fos protein and mRNA levels. It has been shown that ROS increases in AP-1 binding by activating c-jun N-terminal kinase/stress activated protein kinase (JNK?SAPK) or extracellularly responsive kinases (ERK). Ultimately, the generation of intracellular oxidative stress can disrupt normal development by "inappropriately" activating signal transduction cascades to induce the transcription of genese that affect cell growth and differentiation. The hypothesis that cadmium, copper, PCB128 or endosulfan increase the intracellular levels of ROS to affect c-jun levels, and increase AP-1 binding activity will be investigated. To explore this hypothesis two model systems will be used: cultured mammalian cells and zebrafish. The studies proposed using cultured cells will investigate molecular and cellular aspects of hazardous chemical exposure on development. As a physiologic end-point, the effects of cadmium and copper exposure on zebrafish development will be examined.
There are four Specific Aims in this project:
1. Characterize the effects of metals and organic chemicals on metallothionein and proto-oncogene expression
2. Identify upstream regulatory elements that mediate chemically induced transcription
3. Determine the effects of hazardous chemical exposure on the activities of (a) AP-1 and (b) signal transduction cascades that regulate c-jun binding.
4. Evaluate the effects of transition metal-exposure on zebrafish development and signal transduction pathways that regualte development.
The research outlined in theis project is directly relevant to several of the major goals of the Duke University Center. Specifically, this project will elucidate mechanisms of reproductive and developmental toxicicty of selected chemicals that are found at Superfund sites. Furthermore, we will transfer the information obtained using cell culture and zebrafish models to address issues of human and ecological health. The generation of transgenic zebrafish, and subsequent analysis of their response to copper and cadmium exposure, will contribute to our understanding of how transition metals impact reproduction and development in vertebrates. This knowledge will help in the generation of markers for reproductive and developmental toxicities in humans.