Date of Award

2019

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Mary C. Mullins

Second Advisor

Eileen M. Shore

Abstract

Fibrodysplasia ossificans progressiva (FOP) is a rare human genetic disorder characterized by skeletal malformations and progressive extraskeletal ossification. All cases of FOP are caused by activating mutations in the GS or kinase domains of the type I BMP/TGFβ cell surface receptor, ACVR1, which over-activate signaling through phospho-Smad1/5 (pSmad1/5). To investigate the uncertain mechanism by which FOP-ACVR1 enhances pSmad1/5 activation, we used a zebrafish embryonic dorsoventral (DV) patterning assay for BMP signaling. In this system, misexpression of human ACVR1-R206H causes increased pSmad1/5 signaling and ventralization of zebrafish embryos. To pattern the zebrafish DV axis, Acvr1l, the zebrafish homologue of human ACVR1, signals in response to BMP ligand within a heterotetrameric, type I and II receptor complex. Previous studies have demonstrated that type II BMP receptors are required for the activity of FOP-ACVR1, however the roles of ligand and other type I BMP receptors are less well understood. We determined that the FOP mutants ACVR1-R206H and -G328R do not require their ligand binding domain to over-activate BMP signaling in DV patterning. However, intact ACVR1-R206H and G328R have the ability to respond to BMP ligand. Additionally, BMPR1, a type I BMP receptor which is normally required for BMP-mediated patterning of the embryo, is dispensable for both ligand-independent and ligand-responsive BMP signaling activation by ACVR1-R206H. Both ACVR1-R206H and -G328R require a GS domain, and presumptive type II BMP receptor activity. However, ACVR1-R206H and -G328R have a reduced requirement for serine and threonine residues within the GS domain compared to ACVR1. Interestingly, these FOP mutants, differ from each other in their GS domain residue requirements for ligand-independent signaling. Over-active signaling by ACVR1-R206H and -G328R can be abrogated by competitive ATP inhibition, confirming that kinase activity is required for over-active signaling. Lastly, Acvr1l-R203H, the zebrafish homolog of ACVR1-R206H, does not have over-active signaling. However, the human ACVR1 kinase domain is sufficient to facilitate over-active signaling by Acvr1l-R203H. These data provide new insight into how the ligand-receptor complex assembles and is regulated in both wild-type and FOP BMP pathway signaling and elucidate potential targets for the treatment of FOP.

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