In this study, we hypothesize that human acidic mammalian chitinase (AMCase) binds and is regulated by the epidermal growth factor receptor (EGFR), and that AMCase interacts with Galectin-3 (Gal-3) to mediate anti-apoptotic functions. We further hypothesize that asthma-associated polymorphisms of AMCase alter chitinase activity and modulate anti-apoptotic effects. We investigated the interactions between AMCase, Gal-3 and EGFR by establishing binding and co-expression in vitro; apoptotic effects were evaluated via Annexin V/Propidium Iodide staining. Molecular cloning was performed to generate single nucleotide polymorphisms (SNPs) of AMCase associated with asthma. Our data showed that co-expression of AMCase and EGFR induces chitinase activity; we found that AMCase and Gal-3 bind each other in vitro, and that they co-localize in the cytoplasm of cells. Co-transfection of AMCase and Gal-3 demonstrates greater anti-apoptotic effect than Gal-3 alone, while recombinant Gal-3 induces apoptosis, which is not blocked by incubation with recombinant AMCase. From these data, we conclude that AMCase is regulated by EGFR, and that AMCase and Gal-3 physically interact, however contrary to our hypothesis, the anti-apoptotic effects of AMCase are unlikely to be mediated by Gal-3. Further exploration of this pathway using SNP constructs generated in this study will shed light on the mechanism of AMCase in asthma.
| Identifer | oai:union.ndltd.org:YALE_med/oai:ymtdl.med.yale.edu:etd-03102009-162717 |
| Date | 28 September 2009 |
| Creators | Wachtel, Heather |
| Contributors | Jack Elias |
| Publisher | Yale University |
| Source Sets | Yale Medical student MD Thesis |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://ymtdl.med.yale.edu/theses/available/etd-03102009-162717/ |
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