Slowey, Niall (2020) C. elegans Model for Unravelling the Role of Klotho in Ion Metabolism and Aging. Doctoral thesis, University of Huddersfield.

Klotho (KL) and beta-Klotho (KLB) are transmembrane proteins that are cofactors for the endocrine subfamily of fibroblast growth factors (FGFs), including FGF19, -21 and -23. This function means Klotho proteins play important roles in regulating metabolic processes through the activation of FGF receptors (FGFRs). The mechanism by which the Klotho/FGFR complex acts is not fully understood. Klotho also is linked to the regulation of ion channels and therefore ion homeostasis. This project focuses on uncovering the interactions of the Klotho/FGFR complex and other proteins (e.g. ion channels). C. elegans has two orthologues of mammalian KL/KLB, klo-1 and klo-2, two orthologues of FGF ligands, let-756 and egl-17 and a single ortholog of the four FGFR genes, egl-15. Excess activation of EGL-15/FGFR leads to accumulation of fluid in the C. elegans pseudocoelom and a clear phenotype. klo-1 is expressed in the excretory canal, which is responsible for maintenance of fluid balance. A loss of klo-1 or klo-2 function led to the preference to ethanol while the loss of both simultaneously inhibited the preference for ethanol. klo-2 removal was found to cause the over migration of canal associated neurons (CANs) cell body which is essential for excretory canal function. Removal of either klo-1 or klo-2 in aqp-8, (an ortholog of aquaporin water channels), genetic background sensitises the double mutants to environmental changes, whereas simultaneous removal of both klo-1 and klo-2 in aqp-8 background rescues the phenotype. gtl-2 encodes for a TRPM channel protein, which acts in the excretory cell to regulate Mg2+ excretion. gtl-2 mutants show a reduced brood size when exposed to excess Mg2+ which is suppressed by simultaneous genetic removal of both klo-1 and klo-2. Finally, exc-4 encodes for a chloride intracellular channel (CLIC) of anion channels and mutations in exc-4 lead to semi-clear phenotype and the formation pearl structures in the excretory canal. Genetic removal of both klo-1 and klo-2 in exc-4 mutant background significantly rescues the defective pearl structures in the excretory canal. KLO-1 and KLO-2 thus regulate ion homeostasis in synergy with a number of channel proteins, including EXC-4, GTL-2 and AQP-8.

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