Capillary Morphogenesis Gene 2 (CMG2) is a type I transmembrane, integrin-like receptor. It was originally identified as one of several genes upregulated during capillary formation. It was subsequently identified as one of two physiological anthrax toxin receptors, where CMG2 serves as a cell-surface receptor for anthrax toxin and mediates entry of the toxin into cells via clathrin-dependent endocytosis. Additionally, loss-of-function mutations in CMG2 cause the genetic disorder hyaline fibromatosis syndrome (HFS), where the core symptom is dysregulation of extracellular matrix homeostasis (ECM), including excessive accumulation of proteinaceous hyaline material; HFS clearly indicates that CMG2 plays an essential function in ECM homeostasis and repair. Most often, these situational roles have been evaluated as separate intellectual and experimental entities; consequently, whereas details have emerged for each respective situational role, there has been little attempt to synthesize knowledge from each situational role in order to model a holistic map of CMG2 function and mechanism of action in normal physiology.The work presented in this thesis is an example of such a synthesis. Interactions between CMG2 and type IV collagen (Col IV) were evaluated, to better understand this putative interaction and its effect on CMG2 function in angiogenesis. Using an overlapping library peptide array of the Col IV Î±1 and Î±2 chains, it was found that CMG2-binding peptides were enriched within the NC1 domains. This finding was corroborated via another epitope mapping peptide array, where we found a major epitope for CMG2-binding within the Î±2 NC1 domain (canstatin). Identification of CMG2 interactions with Col IV NC1 domains (including canstatin) was both surprising and intriguing, as these domains are potent endogenous inhibitors of angiogenesis. To further evaluate the physiological relevance of interactions with Col IV NC1 domains, a canstatin-derived peptide from the original array was synthesized and used for further studies. This peptide (here known as S16) binds with high affinity (KD = 440 Â± 160 nM) to the extracellular, ligand-binding CMG2 vWA domain; specificity was confirmed through competition studies with anthrax toxin PA, and through demonstration of divalent cation-dependent binding. CMG2 was found to be the relevant endothelial receptor for S16. CMG2 in fact mediates endocytic uptake of peptide S16, as demonstrated by flow cytometry, and colocalization studies. S16 further inhibits migration of endothelial cells. These findings demonstrate that CMG2 is a functional receptor for Col IV NC1 domain fragments. CMG2 may exert a pro-angiogenic effect through endocytosis and clearance of anti-angiogenic NC1 domain fragments. Additionally, this is the first demonstration of CMG2-mediated uptake of an endogenous matrix fragment, and suggests a mechanism by which CMG2 regulates ECM and basement membrane homeostasis, thereby establishing a functional connection between the receptor's role in matrix biology and angiogenesis.
College and Department
Physical and Mathematical Sciences; Chemistry and Biochemistry
BYU ScholarsArchive Citation
Finnell, Jordan Grant, "Anthrax, Matrix Biology, and Angiogenesis: Capillary Morphogenesis Gene 2 Mediates Activity and Uptake of Type IV Collagen-Derived Anti-Angiogenic Peptides" (2017). All Theses and Dissertations. 6849.
angiogenesis, CMG2/ANTXR2, matrix biology, peptide array, type IV collagen