Date of Award

12-2011

Level of Access

Campus-Only Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biomedical Sciences

Advisor

Clarissa Henry

Second Committee Member

Robert Gundersen

Third Committee Member

Mary S. Tyler

Abstract

Extracellular matrix (ECM) remodeling mediates signaling between cells and their microenvironment and is thus critical for tissue development and homeostasis. Matrix metalloproteinases (MMPs) modulate the composition of the ECM by way of degradation of ECM proteins. The activity of these proteases and their tissue inhibitors (TIMPs) must be tightly regulated throughout development and tissue remodeling. MMP misregulation occurs in diseases such as tumor metastasis, arthritis, and fibrosis. The zebrafish myotendinous junction (MTJ), with its localized concentration of multiple ECM proteins and homology to the human tendon, is ideal for studying the in vivo activity of MMPs and TIMPs during matrix remodeling. Two ECM proteins in the zebrafish MTJ are Fibronectin (Fn) and laminin. During MTJ development, Fn is degraded and laminin polymerization increases, however, the mechanisms underlying these dynamic changes in ECM composition are not known. I provide evidence that laminin polymerization is a checkpoint for Mmp11-mediated Fn degradation. Reduced levels of Mmp11 are sufficient to increase Fn levels and ameliorate the disordered myofiber architecture associated with laminin γ1 mutants. Conversely, increased levels of Mmp11 decrease Fn and cause muscle degeneration in wild-type embryos. Although an in vivo inhibitor of Mmp11 was previously unknown, I found that Timp2b affects Mmp11 localization to the MTJ. Decreased levels of Timp2b are sufficient to decrease Fn levels and cause muscle fiber detachment in wild-type embryos. These data provide in vivo evidence of an individual Mmp affecting Fn degradation and suggest that Timp2b is an effector of Mmp11 localization. Both of these findings provide novel insight into the functional roles of individual Mmps and Timps in vivo, which is necessary for developing effective therapeutics against individual proteases. In addition, my finding that adhesion to laminin is a checkpoint for Fn degradation provides greater insight into the understanding of in vivo microenvironment dynamics and the regulation of ECM macromolecules during development.

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