Components of Caenorhabditis elegans cuticular biosynthesis and the molecular basis of morphological mutants

Thein, Melanie C. (2005) Components of Caenorhabditis elegans cuticular biosynthesis and the molecular basis of morphological mutants. PhD thesis, University of Glasgow.

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The cuticle of C. elegans is an extracellular matrix primarily composed of highly processed collagens. It is the site of interaction with the environment, enables motility and controls morphology. Animals with aberrant cuticles have altered body shapes, including dumpiness and a twisted body shape or, exhibit blisters. During the life cycle, C. elegans progresses through 4 moults during which a new cuticle is synthesised and the old cuticle is shed. Synthesis of a new multi-layered cuticle occurs via the temporally-controlled secretion of collagens. The processes by which secreted C. elegans collagens are incorporated into the cuticle are believed to be similar to those of vertebrate collagens, which are co-translationally modified, folded into triple helices, and subsequently proteolytically cleaved at the C- and N- termini. In the C. elegans cuticle, such processed collagen trimers are highly cross-linked by an array of tyrosine-, glutamate, and cysteine-derived inter-chain cross-links in order to produce a cuticle with flexibility and high tensile strength. A C. elegans enzyme, BLI-3, originally termed Duoxl on the basis of the presence of a NADPH-oxidase and a peroxidase domain, has been identified as being an important catalyst of tyrosine cross-links in the cuticle. The H2O2 required for the BLI-3 mediated cross-linking activity is supplied by the NADPH-oxidase domain. A COL-19::GFP marker strain, TP12, has been created and has enabled the visualisation of the tagged collagen in regions and substructures of the cuticle including the annulae, seam cell-derived cuticle and the alae. By creating a number of crossed TP12 strains, this marker has been a useful tool in the analysis of the cuticle structure of many morphological mutants. These studies have enabled the roles of different collagens to be established and also have demonstrated the different mutabilities of discrete cuticle substructures such as the struts, fibrous layers, annular furrows and seam cell-derived cuticle. TP12 has also functioned as a sensitive marker for cuticle disruptions resulting from RNAi-treatment of a number of genes and has shown these disruptions even in the absence of a gross morphological defect. This has identified at least one gene that has a role in cuticle synthesis that previous RNAi screens classified as being wild type. The potential of HPX-1, a putative peroxidase, to mediate cross-linking functions within the cuticle, has been investigated. The RNAi-induced blistered and body morphology defects, as well as its expression being hypodermal and coincident with the moult, are consistent with a role in cuticle synthesis. The Mlt phenotype, which is especially apparent via SEM analysis, additionally implicates HPX-1 in being involved in the moulting process. The relationship between BLI-3 and HPX-1 has also been investigated because BLI-3 possibly represents the sole source of H2O2 for HPX-1 mediated activity. HPX-1 is closely related to peroxidases in other organisms and it has been suggested, based on the roles of these homologues and the observations from these studies, that HPX-1 has synthetic and possibly degradative roles within the C. elegans cuticle.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Microbiology, genetics.
Colleges/Schools: College of Medical Veterinary and Life Sciences
Supervisor's Name: Page, Prof. Tony
Date of Award: 2005
Depositing User: Enlighten Team
Unique ID: glathesis:2005-71192
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 10 May 2019 10:49
Last Modified: 06 Aug 2021 13:46

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