Engineering Insulin: A Manipulative Study on the Activating and Inhibiting Properties of Insulin/Insulin-Like Peptides (INS) in Caenorhabditis elegans
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Authors
Low, Mun-Seong
Date
Type
thesis
Language
eng
Keyword
Caenorhabditis elegans , insulin , modification , INS , worm , IIS , DAF , PTEN
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Abstract
The insulin/insulin-like growth factor signalling (IIS) pathway is evolutionarily conserved between humans and the nematode, Caenorhabditis elegans. While humans only encode activating insulins, C. elegans remarkably encode both activating and inhibiting insulin/insulin-like peptides (INS). Furthermore, strongly activating C. elegans INS contain an F-peptide, where previous work suggested that it may be required for activating activity in certain insulin signalling contexts. However, much is still unknown as to what differentiates an activating insulin from an inhibitor. Previously, the Chin-Sang lab categorized all 40 INS coding genes of C. elegans in relation to their activity on the IIS pathway. Amino acid sequence alignments between the human insulin/insulin-like growth factors and the strongest activators of the C. elegans INS superfamily reveal conserved Glycine residues adjacent to the B-chain/domain disulfide Cysteines. Interestingly, these Glycine residues are not conserved in inhibiting C. elegans INS. I hypothesize that these conserved Glycine residues are vital for the activating properties of INS. To determine this, I overexpressed multiple insulin-like variants and observed their effects on phenotypes associated with abnormal IIS signalling. I report that the B-domain disulfide adjacent Glycine residues are important for activating activity and that I can change a strong activator, like INS-4, to an inhibitor by changing the Glycine residues to Proline. I provide further evidence for the importance of these Glycine residues in conferring activating properties as I could change a strong inhibitor like INS-37 to function as an activator by changing its B-domain disulfide adjacent residues to Glycine. I report that the F-peptide has activating properties and can function as a modulatory activator in F-peptide deficient INS but is specific in only promoting Q-cell divisions. My study is first to show that changing the conserved disulfide adjacent residues is sufficient to alter INS behaviour on the IIS pathway. My work will provide vital information for medical applications in treating insulin-related oncogenic and neurodegenerative disorders as well as progress the current knowledge on insulin/insulin-like peptide properties.
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ProQuest PhD and Master's Theses International Dissemination Agreement
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This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.