Oomatia, Amin; (2024) Investigating the genetic and epigenetic architecture of Mesoamerican nephropathy. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Mesoamerican nephropathy (MeN) is a leading cause of death amongst working-age men in many Central American countries. Renal biopsy data suggests the disease is caused by a chronic tubulointerstitial nephritis, and numerous environmental agents such as heat, heavy metals and pesticides have been proposed to have a causal role in its pathogenesis, yet to date, its underlying cause remains a mystery. A genome-wide association study performed using DNA from the Colt cohort, a longitudinal study based in Nicaragua of individuals at high risk of MeN, has identified two loci associated with the disease: rs4980833 and rs114524020. In silico analysis suggests that rs114524020 represents a CTCF-RAD21 binding site which regulates the expression of a nearby gene: Methylmalonyl CoA Mutase (MMUT), the deficiency of which causes a hereditary chronic tubulointerstitial nephritis. To test the hypothesis that carriage of the minor allele at rs114524020 diminishes MMUT expression in proximal tubules, the locus was knocked out in HK-2 cells using CRISPR-Cas9. qPCR, allele-specific digital PCR and RNA-Seq showed MMUT did not differ between heterozygote rs114524020 knockout cells and wild-type cells suggesting that this locus does not regulate MMUT expression in this cell line. DNA from the Colt cohort was also used to study whole blood DNA methylation as a biomarker for temporally related environmental exposures implicated in the pathogenesis of MeN. Despite having adequate power, there was no suggestion of reported methylation signatures associated with heavy metals, pesticides and ambient temperature, associating with incident disease, providing evidence against their involvement in the pathogenesis of MeN. The work done in this thesis has helped identify potential genetic risk factors of MeN and used epigenetic data as a biomarker to provide evidence against a causative role of heavy metals, pesticides and temperature in the pathogenesis of the disease. Future work could include investigating the effects of rs114524020 knockout in other cell lines and types, and performing a GWAS and EWAS in a second cohort to replicate the second identified risk polymorphism (rs4980833) and DNA methylation signatures reported in this thesis, followed by relevant functional in vitro investigations.

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