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Functional analysis of ATP binding cassette (ABC) transporters in Caenorhabditis Elegans

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(Thesis) Ph.D.
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This thesis deals with functional analysis of ABC transporter genes in Caenorhabditis elegans. ABC transporters constitute one of the largest gene families in C. elegans. Sixty ABC genes have been identified and classified into eight different subfamilies. Fifty-seven orthologous pairs can be found between C. elegans and C. briggsae. As an initial step toward understanding the functions of the ABC genes, I generated transgenic animals containing prornoter::GFP/DsRed fusion constructs for the whole family in order to address when and where these genes are turned on in vivo. I observed frequent expression of ABC genes in intestine, pharynx and excretory cell, especially for those in subfamilies B and C. Sixteen of the 60 ABC genes are tandemly duplicated genes, forming two 4-gene and four 2-gene clusters. Interestingly, the majority of the promoters from the same clusters drive temporally andfor spatially differential expressions, suggesting active function of these duplicated genes in different tissues or stages rather than their being inactivated, as is typical for duplicated genes. Currently 18 single ABC gene mutants have been created. All of them are wild type under normal laboratory conditions. I used one of the ABC genes, pgp-12, an excretory cell-expressing gene, to characterize its transcriptional regulation. A 10 base pair (bp) cis-element, Ex-1 and its binding protein C26C6.5a, were isolated. C26C6.5a is also expressed in the excretory cell and binds Ex-l in vitro. RNAi analysis revealed that the C26C6.5a protein functions as a trans-activator for excretory cell specific expression. The only member of subfamily E is the ABCE gene that has been annotated as an RNase L inhibitor. This annotation is unlikely to be correct because of the lack of the inhibitor gene in C. elegans. Functional analysis of the gene reveals that it is essential for life, possibly involved in transcription and translation, and may function as a nucleocytoplasrnic transporter. RNAi data implies that the protein functions in both germline and vulvae.
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