Investigations into the mechanisms behind human imprinting and imprinting related disorders in model systems

Abstract

Genomic imprinting plays a vital role in normal mammalian growth and development. Normal expression of imprinted genes is highly dependent on parental-specific methylation at the differentially-methylated regions known as imprinting control regions. Imprinting control regions often regulate the parental-specific expression of a number of nearby imprinted genes. Abnormal expression of these imprints is caused by chromosomal anomaly or an epimutation such as loss of methylation at the imprinted control region. Using human and mouse model systems I investigated the mechanisms behind imprinting and imprinting- like disorders.
Firstly, I utilised the chromosomally stable human fibroblast cell line hTERT-1604 with a longterm depletion of the maintenance methylation enzymes DNMT1 and UHRF1. I present data showing that the long-term knockdown of DNMT1 results in an interesting cross-talk between the gametic and somatic DMRs at a subset of imprinted clusters. Further to this, I show the depletion of UHRF1 results in loss of imprinting with the inability to recover upon the rescue of the WT protein.
I also took the opportunity to use publicly available datasets and a bespoke in-house Galaxy workflow to score abnormal methylation variability across patient samples and described a methylation index to detect epimutations in imprinting disorders. Not only was it able to detect the same epimutations as the wet-lab technique that was previously used to diagnose the patient samples, but the methylation index also detected epimutations in patient samples that were not previously diagnosed at the molecular level.
​​​​​​​Finally, with the help of CRISPR-generated mouse models, I was able to describe a novel role for the PHD domain of Uhrf1 in the maintenance of genomic imprinting during embryonic development. Furthermore, I also contributed to the literature in support of Uhrf1’s role in the repression of transposable elements.

Date of Award	Dec 2020
Original language	English
Supervisor	Colum Walsh (Supervisor) & Declan Mc Kenna (Supervisor)