Document Type

Honors Thesis

Major

Biochemistry, Molecular & Cellular Biology

Advisor(s)

Benjamin King

Committee Members

Clarissa Henry, Melissa Ladenheim, Sally Molloy, Kristy Townsend

Graduation Year

May 2020

Publication Date

Spring 5-2020

Abstract

Muscular Dystrophy (MD) is characterized by varying severity and time-of-onset by individuals afflicted with the same forms of MD, a phenomenon that is not well understood. MD affects 250,000 individuals in the United States and is characterized by mutations in the dystroglycan complex. gmppb encodes an enzyme that glycosylates dystroglycan, making it functionally active; thus, mutations in gmppb cause dystroglycanopathic MD1 . The zebrafish (Danio rerio) is a powerful vertebrate model for musculoskeletal development and disease. Like human patients, gmppb mutant zebrafish present both mild and severe phenotypes. In order to understand the molecular mechanisms involved, we performed high-throughput RNA Sequencing (RNA-Seq) and small RNA Sequencing at 4 and 7 days-post-fertilization (dpf) in mild and severe gmppb mutants and controls. We hypothesize that variable phenotypes in gmppb mutants are due to differences in gene regulation; therefore, we identified differentially expressed (DE) long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) - both potent genetic regulators. We identified “MD-relevant” DE Ensembl-annotated genes involved in cell cycle regulation, the immune response, neural development and maturation, and skeletal muscle atrophy. We identified DE miRNAs that regulate these DE genes in the 4dpf severe mutants – identifying 55 of these interactions. We utilized a novel method of visualizing gene expression networks by generating co-expression networks of miRNAs and subsequently removing miRNA nodes to identify important miRNAs. We identified 95 potential lncRNAs for further analysis. By integrating analyses of both coding and non-coding genes, we contributed towards the understanding of the molecular mechanisms of Dystroglycanopathy, highlighting potential phenotypic modulators.

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