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The Known Function and Enigmatic Roles of the Glycine Cleavage System in Development

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posted on 2024-05-09, 16:46 authored by Nicole Elizabeth Weaver
Nonketotic hyperglycinemia (NKH) is a devastating neurometabolic disease that affects every 1 in 76,000 people worldwide. NKH is caused by an inborn error in the glycine cleavage system (GCS), a four-protein complex that functions to break down glycine, methylate folate, and produce one-carbon units to be utilized in other metabolic pathways. Patients suffer from a wide variety of symptoms including seizures, musculoskeletal defects, brain patterning abnormalities, and developmental deviations in other organ systems, including the kidney. Mutations in glycine decarboxylase (GLDC)and aminomethyltransferase (AMT) are responsible for ~80% and ~20% of NKH cases, respectively. Gldc and Amt deficient mouse models have connected the GCS to neural tube and brain development, however, the role of these proteins in kidney development is unresolved. While mouse models are highly genetically similar to humans, there are drawbacks to mice in developmental studies, such as small litters and in utero fertilization. Here, we have employed the zebrafish as an alternative model to study nephrogenesis in NKH, as zebrafish share ~70% of genes with humans, develop ex utero, and generate hundreds of embryos at a time. We have created two novel genetic loss of function models of NKH by targeting gldc and amt. In each NKH zebrafish model, we found decreased survivability, cartilaginous abnormalities, and brain patterning irregularities, recapitulating what has been noted in human patients. Further, both disease models displayed altered fluid homeostasis due to compromised renal clearance. We investigated the effects of glycine accumulation through exogenous treatment, where we discovered similar alterations in nephron patterning to the gldc morphant model. Additionally, all phenotypes were dose dependent, suggesting severity corresponds to amount of glycine that has accumulated. amt models exhibited some unique phenotypes, including a tail kink phenotype and cloacal cysts. We conducted a novel assay to analyze cloacal function through the addition of a fluorescent molecule, NaGreen. We collected live videos of amt deficient animals where we observed a blind ending lumen and an inability to excrete. To probe the genetic cascade responsible for these phenotypes, we utilized a BMP antagonist as BMP signaling has been implicated in cloacal and tail development. We discovered a novel connection between BMP signaling and amt, suggesting dysregulation of amt causes an increase in BMP signaling. Here, we uncovered a new relationship between the GCS, nephrogenic specification, and cloacal development. With the continued use of these models, we will gain a better understanding of the pathophysiology of NKH, which can provide avenues for advancement in therapeutics.

History

Date Created

2024-04-13

Date Modified

2024-05-08

Defense Date

2024-04-05

CIP Code

  • 26.0101

Research Director(s)

Rebecca Wingert

Committee Members

Katrina Adams|Ana Flores Mireles|Molly Scheel

Degree

  • Doctor of Philosophy

Degree Level

  • Doctoral Dissertation

Language

  • English

Library Record

006584478

OCLC Number

1433023876

Publisher

University of Notre Dame

Program Name

  • Biological Sciences

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