View Project rand-affy-droso-366220

Project Summary
Status:	Public
Publications:	1 Published
Project Detail		Data Detail
Platform:	Affymetrix	MIAME Areas	Compliance
Species:	Drosophila	Array Design Detail	true
Organ/Tissue Type:	Late Embryonic tissue	Experiment Detail	true
Organ Region:	Whole brain	Sample Detail	false
Cell Type:	mixed neural cells	Hybridization Detail	false
Study Type:	pharmacogenomic	Measurement Detail	false
Disease/Condition:	normal
Replicates:	3
Expected Samples:

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Investigator Contact Detail
Name	Dr. Matthew D Rand
Institution:	University of Vermont
Street Address:	HSRF 426C
	149 Beaumont Ave
City, State/Province:	Burlington , VT
Zip/Postal Code:	05405
Country:	United States
Work Phone:	802-656-0405
Fax:	802-656-4674
E-mail:	mdrand@zoo.uvm.edu

Proposal Detail

Grant:

ES013754-01

Status:

Public

Service Type:

Start to Finish Profiling

Study Relevance:

Methylmercury (MeHg) toxicity in humans manifests deficits in neurological function. Cases of prenatal exposure to mercury have established that the developing nervous system is most highly susceptible to perturbation by MeHg. At a cellular level, MeHg-induced defects result from altered neuronal proliferation, migration and pathfinding. However, the molecular targets of MeHg that give rise to these outcomes are not fully understood. In an overall effort to identify the fundamental molecular targets of MeHg in neural development, we are investigating the effects of MeHg on gene expression and protein function in the Drosophila model. Since the fundamental signaling pathways in development of multicellular animals have been extensively characterized in Drosophila, and demonstrate high degree of conservation in vertebrates, we believe these data will lead us to the most pertinent pathways affected by MeHg and begin to elucidate the mechanism of MeHg neural toxicity relevant to cases of human exposure to this prevalent environmental toxin.

Hypothesis:

Our hypothesis is that MeHg, by direct interaction with cysteine thiol groups, alters the function signaling pathway proteins and subsequently alters transcription of target genes in the respective pathways. Our hypothesis is supported by our recent data demonstrating a direct action of MeHg in activating the Notch receptor pathway and upregulating target gene expression. We anticipate a microarray analysis will elucidate additional fundamental signaling pathways where transcription is affected by this toxin.

Specific Aim:

Our aim is to identify fundamental signaling pathways in neural development that are targets for MeHg poisoning.

Experimental Procedure and Design:

Fertilized female flies are fed on food containing methylmercury (1-20 micromolar) or solvent control (DMSO) for a period of five days to allow for MeHg penetration in to eggs. Flies are then transfered to a cage for embryo collection for a discrete window of time (e.g. 1hr). Embryos are aged 16-24 hours and collected for total RNA extraction using the Trizol reagent. Total RNA is quantitated by spectrophotometry with a Nanodrop reader. While concentration dependent effects of MeHg are of interest, initial experiments will be conducted on samples exposed to a single concetration known to illicit effect in other bioassays (e.g. 10 micromolar). The goal is to see which genes are up- or down-regulated with MeHg exposure, as compared to control embryos that are not exposed. Emphasis in the analysis stage will be in identifying target genes of known signaling pathways.

Replicate Description:

Biological replicates are obtained from separate batches of female flies fed on MeHg containing food.

Replicate Types:

biological_replicate

Experimental Factors:

Conditions that are tested in the experiment. At least one is required. Experimental factors are the independent variables in the experiment.

Factor Name	Description	Factor Category
Methylmercury	Methylmercury is an organic mercury c...	pharmacogenomic_treatment

Project Samples

Samples associated with this project.

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Name	Description	Extracts
DMSO 1	embryos of flies raised on ...	1
DMSO2	embryos of flies raised on ...	1
DMSO3	embryos of flies raised on ...	1
MeHg 1	Embryos from MeHg fed adults	1
MeHg2	Embryos from MeHg fed adults	1
MeHg3	Embryos from MeHg fed adults	1

Project Hybridizations

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Name	Array	Labeled Extract
drosophila_1_hyb	drosophila_1	DMSO 1_le1
drosophila_2_hyb	drosophila_2	DMSO2_le1
drosophila_3_hyb	drosophila_3	DMSO3_le1
drosophila_4_hyb	drosophila_4	MeHg 1_le1
drosophila_5_hyb	drosophila_5	MeHg2_le1
drosophila_6_hyb	drosophila_6	MeHg3_le1

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