Date of Award

2023-05-01

Degree Name

Master of Science

Department

Biological Sciences

Advisor(s)

Manuel Miranda-Arango

Abstract

The study of glycine transporters has been neglected compared to other members of the Solute Carrier 6 (SLC6) family of proteins, such as the dopamine and serotonin transporters, even though they are just as crucial to survival. However, glycinergic neurotransmission has been extensively studied in mammals, resulting in vast, valuable information. The same cannot be said for invertebrates, however. In this study, we aim to establish an initial first step in elucidating components of glycinergic neurotransmission in the invertebrate, Drosophila melanogaster. We believe this organism possesses a glycine transporter because a glycine receptor has been identified and characterized. Specifically, we hypothesize the candidate gene, CG10804, is a potential gene that encodes for a glycine transporter in Drosophila. We used a bioinformatics approach to determine any conserved residues essential for glycine, sodium, and chloride binding between our candidate gene and its mammalian counterpart. Various uncharacterized D. melanogaster genes were analyzed by homology analysis, and it was determined CG10804 was the most likely candidate to encode for a glycine transporter based on the number of conserved residues with mammalian glycine transporters. Based on the atomic coordinates of the human GlyT1 crystal structure, the amino acids that participate in glycine, sodium and chloride binding were found to be identical to the corresponding residues in the CG10804 protein. Furthermore, gene expression, immunofluorescence and western blot analyses seemingly demonstrate that CG10804 is localized at the plasma membrane of Porcine Aortic Endothelial (PAE) cells, which fulfills a requirement as a member of the SLC6 family. However, uptake experiments demonstrated an insignificant amount of glycine transport from CG10804, suggesting this may not be a glycine transporter in D. melanogaster. However, future studies aim to utilize an insect cell line instead of the mammalian PAE cells, before definitively ruling out CG10804 as the Drosophila glycine transporter.

Language

en

Provenance

Recieved from ProQuest

File Size

p.

File Format

application/pdf

Rights Holder

Denise Avalos

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