Date of Award

12-2006

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Biochemistry and Cellular and Molecular Biology

Major Professor

Ranjan Ganguly

Committee Members

Bruce McKee, John Koontz, Albrecht von Arnim, Sundar Venkatachalam

Abstract

Cytochrome P450 monooxygenases (CYPs), are involved in the metabolism of a diverse group of endogenous and xenobiotic compounds. In insects, CYPs are involved in conferring resistance against insecticides. In Drosophila, the expression of Cyp6a2, Cyp6a8, Cyp6g1, Cyp6w1 and Cyp12d1 is higher in the resistant compared to susceptible strains. Recent study by Daborn et al (2002, Science, 297, 2253-2256) showed that Cyp6g1 alone can confer resistance phenotype. The aim of my first objective was to re- examine this claim and second to examine the effect of common xenobiotic compounds on the transcriptome of Drosophila.

In first objective, six strains of Drosophila were examined for DDT resistance and Cyp6g1 expression. Results showed that some of the highly susceptible strains showed high level of Cyp6g1 expression and Accord element in the Cyp6g1 upstream DNA. When Cyp6g1 allele of the resistant 91-R strain was substituted with that of the susceptible 91-C strain via recombination, the resulting three recombinant lines retained high level of resistance like the 91-R strain, but showed very low Cyp6g1 expression. This suggests that there is a correlation between overexpression of Cyp6g1 and the presence of Accord transposable element but not DDT resistance.

In the second part of the first objective, I directly examined the role of the Cyp6a2 and Cyp6g1 in DDT resistance. Germ line transformation in susceptible strain showed that there was a two-fold increase in DDT resistance (LD50) in transformed flies showing two-fold higher expression of GAL4/UAS driven CYP6A2 or CYP6G1 cDNA. A cumulative increase (4-fold) in DDT resistance was observed when both cDNAs were overexpressed in the same fly. Results suggest that the expression of multiple Cyp genes may be needed to confer a high level of DDT resistance.

In the second objective, microarray was used to examine the transcripts induced by caffeine and phenobarbital. Results showed that genes involved in detoxification, carbohydrate metabolism, signal transduction and Cyp genes are induced by caffeine and phenobarbital. These are the same group of genes overexpressed in the resistant 91-R and recombinant strains. These studies shed light on the molecular basis of induction of Cyp genes and insecticide resistance.

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