Date of Award
Doctor of Philosophy (PhD)
Robert O. Heuckeroth
ABSTRACTROLE OF ACTG2 MUTATIONS IN VISCERAL MYOPATHY Sohaib K. Hashmi Robert O. Heuckeroth
Visceral myopathy is a debilitating condition characterized by dysfunction and weakness of smooth muscle in visceral organs including bowel, bladder, and uterus. When the bowel is primarily affected, the disease is called myopathic chronic intestinal pseudo-obstruction (CIPO). Multi-organ dysfunction involving bowel and bladder is known as Megacystis Microcolon Intestinal Hypoperistalsis Syndrome (MMIHS). Individuals affected by CIPO/MMIHS experience feeding difficulties, growth failure, life-threatening abdominal distension, and become dependent on intravenous nutrition for survival. Unfortunately, due to our limited understanding of the pathophysiology of visceral myopathy, current therapies are only supportive, with no mechanism-based treatments available. The most common genetic causes of CIPO/MMIHS are mutations in gamma smooth muscle actin (actin gamma 2, smooth muscle; ACTG2). Disease-causing ACTG2 mutations result from a single amino acid change in one allele (heterozygous missense mutations), with Arginine 257 to Cysteine (R257C) being the most common mutation. To understand why this ACTG2 mutation causes disease, I developed a framework to understand the pathophysiology of ACTG2 -related visceral myopathy. First, I engineered novel tools to quantitatively study the effects of ACTG2 R257C on organization of actin filament bundles and actin-mediated cellular functions in cultured primary human intestinal smooth muscle cells (HISMCs) (Chapter 2). I discovered that the ACTG2 R257C mutation led to fewer, shorter, thinner, and less branched ACTG2 filament bundles without affecting global actin filament bundle organization in HISMCs. I found that ACTG2 R257C-expressing HISMCs spread faster and were more migratory than ACTG2 WT-expressing HISMCs (Chapter 2). I also generated patient-derived and CRISPR gene-edited hPSC lines with the ACTG2 R257C mutation and developed a novel method for differentiating hPSCs into smooth muscle-like cells that express ACTG2 at higher levels than previously achieved (Chapter 3). I then developed a strategy and tools to generate the first Actg2 in vivo disease model, a conditional Actg2 R257C knock-in mouse, for which our efforts are ongoing (Chapter 4). My approaches are modular, facilitating studies of other disease-causing ACTG2 mutations, and enhancing our understanding of ACTG2 disease pathophysiology, which we hope will lead to new treatments.
Hashmi, Sohaib Khalid, "Role Of Actg2 Mutations In Visceral Myopathy" (2020). Publicly Accessible Penn Dissertations. 4647.