Date of Award
Summer 2024
Document Type
Thesis
Degree Name
Master of Science (MS) in Biology
Department
Biology
Committee Chair
Ashley Turner
Abstract
Polycystic ovary syndrome (PCOS) is a complex disorder with various implications, such as polycystic ovaries, visceral obesity, and increased risk of cancer. YAP1 was recently identified as a gene of interest in the development of PCOS. Researchers have established that single nucleotide variants in YAP1 are likely to play a role in PCOS development. This project aims to provide insight into the potential impact of a YAP1 variant of uncertain significance (VUS). Studies in C. elegans have established yap-1 as a nematode ortholog for human YAP1. A YAP1 VUS was identified through ClinVar, YAP1 c.1015A>G (p.Asn339Asp). Evolutionary conservation of the VUS loci was confirmed using multiple sequence alignments in Benchling. Additional analyses were carried out, examining the difference in amino acid chemical properties and the bioinformatics PolyPhen-2 prediction. PolyPhen-2 predicts this variant to be likely pathogenic (HumDiv score of 0.982). These findings supported further investigation, and we sought out to examine the VUS in vivo through the C. elegans yap-1 ortholog and model. DNA primers were designed and optimized using PCR to amplify the YAP1 VUS loci in the nematode yap-1 region. A CRISPR RNA guide was designed and optimized to target yap-1 using an in vitro nuclease assay. The VUS-yap-1 C. elegans strain was generated through microinjection of CRISPR-Cas9. The generated VUS strain will be assessed for possible phenotypes. This study has provided insight into the potential impact and significance of a patient VUS through in silico and in vivo studies. Future bioinformatic experimentation will include protein modeling to examine potential structural changes between encoded wildtype and mutant proteins.
Included in
Bioinformatics Commons, Genetics Commons, Medical Genetics Commons, Molecular Genetics Commons